FACTORS AFFECTING THE ROOT- REGENERATING POTENTIAL OF TAXUS Thesis for the Degree of M. S. MICHIGAN STATE UNIVERSITY JUDITH K. LATHROP 1969 (E, LIBR AR, Michigan 31:. Univcn‘ty III III II III IIIIIIIIIIIIIIIIIIIIIIIIIIIIIII I/ 3 1293 10437 7662 , -. s. alumna IV 1- " I TIME 8 SUIS' I I‘ TBESAEIEDWY mc . i.[ all.-- IIDITITLII‘ ABSTRACT FACTORS AFFECTING THE ROOT-REGENERATING POTENTIAL OF TAXUS By Judith K. Lathrop The yearly cycle of the root-regenerating potential (RRP) of three-year-old Taxus hunnewelliana, as measured by the number of new root initials produced six weeks after lifting, displayed a distinct periodicity. During the summer months (June 21 to August 30) very few active root initials were produced. The RRP increased in the fall (September 13 to October 14), followed by continued increases in the late fall and early winter (October 14 to December 9). A peak period in the RRP occurred in mid~winter (December 9 to February 16). This was followed by a sharp, pro- longed drop in the RRP from March 1 to June 7. Dormant root tips of Taxus cuspidata are characterized by a dark red root tip and an internally metacutized endodermis and root cap connected by metacutized cortical cells. Root dormancy of one-year—old Taxus cuspidata is broken after exposure to seven Judith K. Lathrop weeks of chilling in the dark at 35° F. ; ten weeks of chilling at 28C, , 320 and 40° F. ; and after twelve weeks of chilling at 48° . The resultant shoot growth, however, significantly increases after three weeks of chilling at 28° and 32° F. ; four weeks of chilling at 35° and 40° F; and five weeks of chilling at 48° F. After the chilling require- ment for shoot growth has been fulfilled, daylength exerts no sig- nificant effect on percent buds broken and length of new shoot growth. The cold requirement for root dormancy appears to be "sensed" by the roots themselves. Cold treatment to shoots increases root development but does not substitute directly for the chilling requirement of the roots. Judith K. Lathrop Summer, 1969 FACTORS AFFECTING THE ROOT-REGENERATING POTENTIAL OF TAXUS By Judith K. Lathrop A THESIS Submitted to Michigan State University in partial fulfillment of the requirements for the degree of MASTER OF SCIENCE Department of Horticulture 1969 £5727? 7*3—é7 ACKNOWLEDGMENTS I wish to express my sincere thanks and appreciation to Dr. Roy A. Mecklenburg for his guidance and assistance throughout my graduate program and thesis preparation. Special appreciation is expressed‘to Knute E. Miller for his assistance in the statistical analysis of the data; and to Professor Clarence E. Lewis whose moral support and encouragement throughout my graduate program made it a very-worthwhile and pleasant experience. The financial support of the Michigan Division of the Women' s National Farm and Garden Association is gratefully acknowledged. Without the support of their fellowship and the con- tinuing support of a teaching assistantship, my studies at Michigan State University would not have been possible. Judith K. Lathrop Summer, 1969 ii TABLE OF CONTENTS ACKNOWLEDGMENTS LIST OF TABLES LIST OF FIGURES LIST OF APPENDIX TABLES INTRODUCTION LITERATURE REVIEW . Root-Regenerating Potential . Anatomy and Browning of Roots Cold Requirement Shoot Bud Dormancy Root Dormancy. ME THODS AND MATERIALS Root-Regenerating Potential . Anatomy . Cold Requirement . . . One- Year- Old Taxus cuspidata Two- Year- Old Taxus cuspidata Daylength after Chilling. . . Differential Chilling of Root and Shoot RESULTS AND DISCUSSION . Root-Regenerating Potential . Anatomy iii Page ii vi . viii 11 11 16 18 18 19 19 19 21 21 22 24 24 29 Cold Requirement Root Dormancy Shoot Dormancy . . Daylength after Chilling. Differential Chilling of Shoot and Root CONCLUSIONS . LITERATURE CITED . APPENDIX TABLES iv Page 35 35 39 45 51 55 61 66 Table LIST OF TABLES The yearly cycle of the root-regenerating potential of Taxus hunnewelliana. (Total number of new root initials produced 6 weeks after lifting.) Effect of daylength on shoot growth of Taxus cuspidata after 12 weeks exposure to 5 tem- peratures Differential chilling of shoot and root . Page 27 50 52 Figure 10. LIST OF FIGURE S The yearly cycle of the root-regenerating potential of Taxus hunnewelliana . Dormant root tip, intermediate root tip, active root tip of Taxus cuspidata Dormant lateral root tips of Taxus cuspidata Longitudinal section of an active root tip of Taxus cuspidata Longitudinal section of a dormant root tip of Taxus cuspidata Mean percent active roots of one-year-old Taxus cuspidata cuttings, two weeks after temperature treatment Mean percent active roots of two-year—old Taxus cuspidata cuttings, two weeks after temperature treatment One-year-old Taxus cuspidata cuttings exposed to 1 through 4 weeks of chilling at 35° F. , as compared to cuttings which remained in the greenhouse (minimum temperature 65° F. ) Mean length of shoot growth (cm.) of one- year-old Taxus cuspidata cuttings, twelve weeks after temperature treatment Mean percent buds broken of one-year-old Taxus cuspidata cuttings, twelve weeks after temperature treatment vi Page 26 31 31 33 33 36 37 41 42 43 Figure 11. 12. Page Mean length of shoot growth (cm. , six replications) of one-year-old Taxus cuspidata cuttings exposed to 2 daylengths after 12 weeks of chilling at5temperatures 47 Mean percent buds broken (six replications) of one-year-old Taxus cuspidata cuttings exposed to 2 daylengths after 12 weeks of chilling at 5 temperatures 49 vii Table 1A. 2A. 3A. 4A. 5A. 6A. 7A. LIST OF APPENDIX TABLES Analysis of variance for overall regression (root-regenerating potential of Taxus hunnewelliana), regression coefficients, multiple correlation coefficient and standard error of estimate Analysis of variance for percent active roots of one-year-old Taxus cuspidata cuttings . Analysis of variance for percent active roots of two-year-old Taxus cuspidata cuttings Analysis of variance for length of shoot growth (cm.) of one-year-old Taxus cuspidata cuttings . Analysis of variance for percent bud break of one-year-old Taxus cuspidata cuttings Analysis of variance for length of shoot growth of Taxus cuspidata exposed to 2 daylengths after 12 weeks of chilling at 5 temperatures . Analysis of variance for percent bud break of Taxus cugfidata exposed to 2 daylengths after 12 weeks of chilling at 5 temperatures . viii Page 66 67 68 69 70 71 72 INTRODUC TION Needle—like evergreen nursery stock (Eggs s22. , Juniperus sap. ) is most frequently propagated by hardwood cuttings taken in the late fall and early winter. After a 2-3 month rooting period in the propagation bench, these cuttings are either transplanted outside or potted and grown under a lath shade structure. The first and possibly the second transplanting is done bare-root. They remain under a lath for one or two years, after which they are lined out in the nursery field row. It is a common practice in Michigan nurseries to dig larger plants with a ball of soil containing the root system to protect the finer roots during transplanting. There is very little chance of successfully transplanting larger evergreen stock if the soil is removed from the root system at the time of digging and it remains in this condition until transplanted. A significant amount of the root system is lost necessarily even in the ball and burlap pro- cess, as only a moderate sized ball of soil is feasible for storing and shipping. Poor survival of transplanted evergreen stock may be due to external environmental factors such as competing vegetation, rodent damage, soil moisture and aeration. However, some of the planting failures may be attributed to the physiological condition of the planting stock (41). As pointed out by Stone (41), the failure of the root system to redevelop in proportion to shoot growth after transplanting is perhaps the most critical factor in determining whether a satisfactory physiological condition exists for transplant- ing. If the root system fails to develop shortly after transplanting, the seedling would probably die because of lack of available soil moisture. The purpose of this investigation is to (1) determine the factors which influence the regeneration of roots on existing root systems of Taxus _s_pp. , (2) examine the interrelationship between shoot growth and root growth during the dormant period, and (3) establish criteria for defining root dormancy as it occurs in Taxus 31E LITERATURE REVIEW Root-Regenerating Potential The survival of bare-root transplanted nursery stock is dependent upon the rapidity with which a new root system is developed. If the transplanted seedling is unable to regenerate a new root system rapidly to tap new sources of moisture, the seed- ling dries out and dies. It has only been in the last fifteen years that published reports dealing with root growth on recently trans- planted seedlings have appeared (Pinus radiata, 15; Pinus ponderosa, 16, 41, 42, 44, 53; Pinus jeffreyi, Abies mflnifica, Abies concolor, 41; Pseudotsuga menziesi, 45). Stone et al. (42) define the ability of a recently transplanted seedling to elongate existing roots or initiate new roots as the root-regenerating potential (RRP). The root-regenerating potential displays a distinct perio- dicity(15, 16, 41, 42, 44, 45, 53). This has been shown quite clearly by Stone and Schubert (42) who evaluated the root regenera- tion of ponderosa pine (Pinus ponderosa) seedlings lifted at monthly intervals throughout one year. The root regeneration of these two-year-old pine seedlings was evaluated 30 days after originally being dug and replanted in the greenhouse at constant root moisture and temperature conditions. The regenerated root system, then, was a result of elongation of existing or remaining short lateral roots and initiation and elongation of new lateral roots. It may be generally stated that there are two periods of active root elongation: one in the spring and one in the fall (16, 22, 41, 42, 44, 45). Some root elongation may occur at all times; however, seedlings lifted in July and August showed very little or no lateral root elongation or root initiation (42). Lateral root elongation on ponderosa pine (Pinus ponderosa, 42) and Douglas-fir (Pseudotsga menziesi, 45) occurred from September to June, reaching maximum amounts from March to May. Lateral root initiation, however, was evi- denced only on seedlings transplanted from December to June, with peak activity occurring in spring prior to terminal bud break. This suggests that elongation and initiation of lateral roots are not con- trolled by the same factors. The endogenous rhythm and/ or envi- ronmental factors which control root elongation are not of the same character or strength as those factors which control root initiation (19). The natural environment before lifting and conditions during the subsequent transplanting period may affect the root-regenerating potential. Optimum soil moisture content and soil temperature are the most critical factors involved in determining the RRP following transplanting (22, 46).. Stone gi_a_l. (45) indicate in preliminary studies that a wide range of photoperiods and variable day tempera- tures during their one month test period has very little effect on the number of roots produced. Therefore, environmental conditions before lifting appear to trigger the cyclic or periodic nature of the RRP. Schubert and Baron (37) and Krugman e_t_al. (16) were the first to report a relationship between root response and exposure to low night temperatures before lifting. Krugman e_tgl. (16) have shown a significant increase in the number of existing roots which elongated after exposure to 90 cold nights (6° C), increasing signifi- cantly up to 150 cold nights. However, the number of newly initiated roots did not significantly increase until exposed to 150 cold nights. After exposure to at least 90 cold nights with accompanying warm, long days, 28% of the ponderosa pine buds expanded. This period of time was sufficient to complete the metabolic changes which pre— cede bud break. Internal metabolic changes were not complete after 90 cold nights to bring about an overall sharp increase in the RRP, but required 120 to 150 cold nights. These differences in shoot/ root cold response suggest two distinct and separate promotor- inhibitor systems are operating. It is also possible that shoot growth after 90 low temperature night treatments confounded the RRP data which occurred after 120-150 cold nights. The comple- tion of the metabolic changes necessary for bud break, before the metabolic changes were complete for root growth, may certainly have an effect on subsequent root growth (i. e. , beyond 90 days). Plants growing in a more natural environment do not experience conditions promoting shoot growth before the cold response stimu- lating the RRP is satisfied. Short photoperiods and cold day tem- peratures throughout the winter and into the spring prevent bud break from occurring even though the cold requirement is satisfied and physiologically the bud is capable of expansion and growth. By this time the roots also have been sufficiently cooled and the meta- bolic potential for root initiation and/or elongation is at a maximum. In the spring, with the onset of longer photoperiods and warmer days, the buds expand with a concomitant decrease in the RRP. The importance of spring planting is well recognized (44). At this time of the year, with early spring rains, soil moisture is at field capacity, accompanied by increasing soil and air tempera- tures. These conditions are very favorable for establishment of transplanted stock. It is economically advantageous to be able to dig such evergreen stock in the fall dormant period (42, 24), and place in cold storage until spring transplanting. Stone and Schubert (43) propose that ga_rly fall lifted stock held in cold storage fails to_ achieve ”physiological hardening" or "readiness" before being lifted. In other words, the seedlings must attain a physiological state, con- ditioned by the natural fall to early winter environment to develop a high RRP which withstands cold storage. The exact environmental conditions necessary to produce this "readiness" and a high RRP have not been determined. These conditions also may vary-with the plant and the precise dates at which plants should be lifted to obtain the best survival in the spring cannot as yet be stated. Anatomy and Browning of Roots A state of dormancy exists in roots of some tree species (33), while others have been reported to have no inherent dormant or rest period (8, l4). Certainly the nature of root dormancy, its causes and control, have not been studied to the same degree as shoot dormancy. Tips of dormant tree roots exhibit unique physio- logical and anatomical characteristics. It has been well documented that dormant or non-growing root tips are often an opaque brown whereas tips of growing roots are often a translucent white (12, 33, 49, 50). Such external changes are interpreted as indicative of anatomical changes occurring within the root (33). Wilcox (49) describes the process, termed metacutization, by which a dormant root tip is formed. Metacutization is a combined chemical process wherein lignification occurs in certain cells of the cortex with simul- taneous suberization throughout these same cortical cells (49). Initial investigations by Plant, as summarized by Romberger (33), describe the common occurrence of the metacutized layer in dormant roots of gymnosperms. Four distinct types of dormant roots occur in gymnosperms according to the degree of metacutization and its relationship to the endodermis (33). The sequence of metacutization as it occurs in noble fir (Abies procera) (49) proceeds in an acropetal direction and follows a regular pattern. As the root tip ceases to elongate, the endodermis differentiates to the region near the apex. The endodermis becomes suberized in its develOpment and this zone of suberization extends out from the tip of the column of endodermis through the cortex to the root cap. The suberized layer continues in the root cap cells until the apical initials and their immediate derivatives are sur- rounded. In root tips of red pine (Pinus resinosa) investigated by Wilcox (50), the final pattern of the metacutized layers is the same as in noble fir; however, the order in which suberization takes place is different. The root cap cells and the endodermis suberize, apparently simultaneously, with the final development of a bridge of metacutized cells extending from the root cap to the suberized endodermis. The brown coloration noted at the tip of dormant roots is due to the degradation of those cells external to the metacutized layer and effectively closed off from the rest of the root. The pat- tern of formation of the metacutized layer in noble fir (49) and red pine (50) and the pattern which occurs in white pine (Pinus strobus) (12), is similar to the type II metacutization pattern found by Plaut (33) to occur in Pseudolarix __p. Browning of the root tip, therefore, is not a causal phe- nomenon of root dormancy but a phenomenon which may or may not accompany metacutization. If the outer most cells of the root cap become metacutized, the root tip may not develop the brown colora- tion associated with a dormant root even though internally the root is structurally dormant (50). This becomes a problem in visual discrimination between active (white) and dormant (brown, red) root tips. .Variations in the production of active and dormant roots may be considered "seasonal" in view of the whole root system and "individual" in view of the differences between roots on the same plant at the same time. At any one time of the year some dormant and some growing roots are present (33, 41, 49, 50), but the ratio of dormant to active roots varies with the season. In coniferous 10 species such as Abies procera (49), Pinus resinosa (22, 50), and others (33), there is a distinct seasonal periodicity in the production of active and dormant roots in field soil. In general, this seasonal variation is characterized by a flush of root growth in the spring, usually before or at bud break, followed by a lull in the summer, with a slight increase in the fall and a prolonged lull in root growth in the winter. During the active growth periods the ratio of dormant to active root tips is low due to a predominance of actively growing roots. The converse may be said of the two periods of relatively little or no root growth in the summer and winter months where the ratio of dormant to active roots is high. The root system is not all dormant or all active at any one time; individual variations do exist (33, 49, 50). Seasonal changes in the length of white unsuberized root on fruit trees (9, 34, 35) and in some coniferous species (13) are depen- dent on the relative rates of root growth and the subsequent rate of cortical browning. All growing roots remain white for only a limited length of time. The outer cortical tissues eventually become suberized and turn brown acropetally, until a non-growing root tip itself browns (9). When the root tip is actively growing and elongat- ing, it has a considerable length of white root; the browning process does not catch up until elongation stops (9, 49, 50). Therefore, the length of white root existing on a root system at any one time is a 11 function not only of the rate of growth but also of the rate of cortical browning or suberization. Head (9) has plotted the seasonal fluctua- tions in cortical browning of fruit tree roots which follows closely the seasonal periodicity in production of actively growing roots. That is, there is an increased rate of cortical browning in the spring which does not keep pace with increased root activity and results in a greater amount of white root. Head (9) reports a lower, relatively constant rate of cortical browning Of roots from late May to Septem- ber; therefore fluctuations in the amount of white root present is a result of new root production and growth. A very slow rate of browning of roots occurs in winter, some roots remaining white for more than three months. White root present in the winter is more likely to be a result of slow rates of suberization of roots rather than active root growth (9, 35). Unless the "periodicity" of cortical browning is established for the test plant, root initiation is perhaps a more accurate and detectable indication of the potential activity of a root system than measures of root elongation by the length or presence of white root. Cold Requirement Shoot Bud Dormancy Smith and Kefford (38) have described an annual pattern of bud development consisting of three major phases. Following the 12 spring flush of growth the three phases are as follows: (1) dormancy development leading to the dormant state; (2) release from dormancy leading to the non-dormant state; and (3) the initiation of spring burst leading again to spring development. The current definition of a dormant bud is one which is unable to grow under conditions of temperature and nutrition normally suitable for growth (32, 36, 38, 48). It has been almost fifty years since the importance of a winter chilling period for breaking the rest period in plants was first recognized (33). Howard (1910) (10) made the first extensive study of the ability of 283 deciduous woody species to sprout when brought into the greenhouse after exposure to various lengths of natural winter environment. Of plants brought in in November, only 50% grew, while 860/8 sprouted from plants brought in in January. Fur- ther experiments with various plant species indicated to Howard (10) that (1) some plants had no definite rest period and were capable of sprouting whenever placed in a warm moist environment and (2) some plants were capable of forming a very profound, deep rest and would not sprout under greenhouse conditions if brought in before late winter. Howard (10) believed that external conditions deter- mined the time of occurrence and degree of intensity of the dormant winter period. Howard (10) had described the need of "resting" by 13 plants in order to undergo internal changes, but it was not until Corville (2) accurately described the period of "resting" as, in fact, a period of chilling that the importance of a Cold requirement was recognized. Corville' s general conclusions are, in large part, still valid today. (1) In cold climates dormancy may set in before cold weather, the onset of complete dormancy not being dependent on cold weather. (2) A period of chilling is necessary before these plants will resume normal growth. (3) Growth of unchilled plants occurs much later in spring than those that have been subjected to a period of chilling. (4) The effects of chilling are limited to those plant parts subjected to the chilling (2). There are also alterations in growth of plants which have not been chilled (1, 2, 36, 48, 52). Time may eventually overcome the dormant state, and growth eventually begins again; but it is irregular and stunted in compari- son with plants receiving their chilling requirement. The economic importance of the chilling requirement for opening of buds of decidu- ous ornamentals and orchard trees is discussed by Chandler et a1. (54). Gustafson (7), working with seedlings of red pine (Pinus resinosa) and white spruce (Picea glauca), noted that the latter (had no chilling requirement and no rest period, whereas red pine does require cold temperatures to break dormancy. Red pine seedlings (7) l4 and sweetgum (Liquidambar'styraciflua) seedlings (6) receiving no chilling, could be forced to grow if exposed to 16 hour photoperiods, although growth in length was somewhat less if not chilled. There- fore, in some cases, the chilling requirement may be overriden by long photoperiods (Picea EEE' , 25, 26; Taxus cuspidata, 39; 6, 7, 33, 36). Temperature and light do not always act independently. Shoot growth stops in many'woody plants when exposed to short days which Downs (4) explains in terms of the phytochrome system. Under long day conditions, many woody plants exhibit continued growth (4). Downs points out that low night temperatures may prevent a long- day growth effect (4). Nienstaedt presents more recent evidence (25, 26) supporting the alternate hypothesisthat white spruce (M M) possess a true period of dormancy maintained by internal conditions of the bud. That is, it is not a "quiescen " condition imposed by an unfavorable environment which can easily be reversed. White sprucedoes have a definite chilling requirement which varies depending on the state of dormancy (or time of year) and the age of the plant (25). The cold requirement of seven spruce species studied (26) maybe overcome by long photoperiods. In studies on Norway spruce (Picea abies), Worrall et a1. (52) have shown that variations in leafing out (bud break) are related to temperature. Both photo- period and chilling cause breakage of true dormancy; however, as 15 length of chilling increases the promoting effects of photoperiod rapidly decline to zero (6, 52). Farmer (6) has shown on sweetgum that some photoperiodic effects occurred at all levels of chilling until the cold requirement was satisfied (1200—1260 hours of chill- ing); after which normal bud break results under any daylength conditions (6, 47). In unchilled plants of Norway spruce (52),. leaf- ing out had not occurred after seven months in an 8 hour photoperiod but had occurred slightly before seven months in a 16 hour photo- period. It appears from this data that daylength is not a major con- trolling factor in breaking winter bud dormancy in Picea abies (52). From the above discussion, it becomes obvious that the effectiveness of photoperiod in breaking bud dormancy varies even among species of the same genus and may not be a major controlling influence. An increase in temperature, not daylength, is the main environmental factor controlling the spring flush of growth once the cold require- ment is satisfied. Samish (36) defines two parameters involved in the chilling concept. (1) A threshold temperature exists below which all tem- peratures have the same effect in breaking rest (17, 33, 48); and (2) a cumulative number of hours of exposure to a temperature below the threshold value is required to terminate rest. In some cases exposure to a warm period before the cold requirement is met may 16 reverse the chilling effect (33, 36, 38). Extended periods of chilling cause no significant adverse effects or promotive effects; that is, a maximum is reached after which further chilling no longer decreases the days till leafing out. Under these conditions, Worrall $11. (52) define the chilling requirement "as that period of chilling at which a further 10—days of chilling produces less than one day acceleration in breakage of dormancy. " Root Dormancy When true dormancy is released (through the cold require- ment) changes occur in the bud even though bud expansion may not immediately follow (32). Winter root grthh in seedlings of Acer saccharinum is controlled by the bud (32). Richardson (32) has shown that root growth will occur only if a physiologically non- dormant bud is present. Bud expansion is not a necessary pre- requisite for stimulating this root activity, and once the chilling requirement for the bud is met it becomes a physiologically non- dormant bud capable of exporting the necessary growth factors to the root (32). Typically, in the field, the chilling requirement is met several weeks before external conditions are favorable for bud expansion. It is during this period of imposed bud dormancy that root growth is possible in the field, assuming of course that soil temperatures are not limiting (32). Active root growth in apple 17 (34, 35), filbert (8), and others (3) occurs throughout the winter when soil conditions are favorable even though above ground portions of the plant are dormant. In mild climates, then, apple and filbert roots continue to grow irrespective of the time of year. Also in coniferous species, such as Taxus media "Hicksii" (Taxus media = Taxus cuspidata X Taxus baccata) root growth is limited by soil temperature and these roots continue to grow when the shoots have become completely dormant if the soil temperature is maintained above 2°C. (23). Root growth in some species is dependent upon conditions which affect the shoot, namely light intensity (5) and air tempera- ture (28, 29, 30, 31, 32), and also upon rOot temperature (8, 20, 27, 29, 51). Root elongation of Acer saccharinum in the non- dormant period is greatly affected by shoot temperature and root temperature (28). Eliasson (5) found a significant depression in root elongation with decreasing light intensities applied to the shoot below 4000 LUX in Populus tremula. Root growth, then, is closely dependent on the rate of photosynthesis (5), the amount of stored carbohydrates in the leaves, stems (5) and in the roots (30) and possibly accessory growth factors (31). METHODS AND MATERIALS Root-Regenerating Potential Three-year-old container—grown Taxus hunnewelliana (Taxus cufiidata X Taxus canadensis) cuttings were used in this study to determine the root-regenerating potential. Field grown plants were dug with intact root systems and grown in containers out-of-doors under a lath shade structure. The containers were plunged in peat moss for the one-year duration of this study. Five plants were brought into the greenhouse at biweekly intervals for a period of one year. All white root was removed and only the woody part of the root system remained after this severe pruning. These plants were immediately planted in a ”turface” (Wyandotte Chemical Co. ) medium, fertilized (completely soluble 20-20-20) and placed in a growth chamber having a constant 60° F. temperature, 10 hour photo- period under approximately 1300 ft. candles. After six weeks each plant was carefully removed from the pot, the turface being gently washed from the root system with water. All new root initials were counted. This is a measure of only root initiation, not root 18 19 elongation, as the regenerated root system was derived from thick woody roots, all short lateral roots having been removed when brought into the greenhouse. Anat omy Dormant roots are easily distinguished in the winter by the presence of a dark red root tip. A great majority, but not all, of the non-growing roots have a dark red root tip. A few roots have a light tan color characteristic of non—growing roots, but do not have a dark red root tip. Samples of actively growing root tips and red, dormant root tips were collected from the root system of M cuspidata. These samples were frozen in liquid nitrogen and stored until sectioning. Longitudinal sections were made on a cryostat and stained with Sudan IV (12). Cold Requirement One -Year-Old Taxus cuspidata To investigate the effects of cold temperatures, one-year~ old rooted cuttings of Taxus cuspidata were exposed to low tempera- tures for a period of one through twelve weeks beginning in early December 1968. These cuttings were grown in a soil-sand mixture in 2:11- " clay pots and had not been exposed to natural fall'or early 20 winter low temperatures. Five groups of plants were placed in cold temperature rooms in the dark maintained at a constant 28° , 32° , 35° , 40° and 48° F. , respectively. Each week four plants from each temperature treatment were removed from the cold temperature rooms and placed on greenhouse benches. After two weeks in the greenhouse at a minimum of 55° F. day—night temperature and normal daylength, measurements were taken of root activity. Each cutting was removed from its pot and the soil care- fully washed from the root system. A continuous segment of root was removed from the root system in such a way as to include the long, thick, extended roots as well as the finer root tips forming a dense mat at the base of the original stern cutting. No fewer than A 30 root tips and/or 20 cm. of root were collected. The number of white, active root tips and non-growing root tips were recorded for each cutting. After sampling the root system, the cuttings were repotted and remained in the greenhouse. Since two weeks was not sufficient to allow for full expression of shoot growth, bud activity was recorded after each group had been in the greenhouse for twelve weeks following cold exposure. Total number of buds, the number of buds that had expanded and length of new shoot growth was assessed for each cutting. 21 T‘wo-Year-Old Taxus cuspidata In a similar manner, the root activity of two-year—old _T_a_x_1_1_§ cuspidata cuttings was studied for a period of eight weeks beginning in late February 1969. These cuttings were potted in a soil-sand mixture and had been exposed to a natural winter environment the winter of 1967-1968. They were moved into the greenhouse at the end of the summer of 1968 in order to prevent exposure to natural cold temperatures in the current dormant season. Daylength after Chilling To study the effects of daylength on shoot growth after cold treatment, one-year-old Taxus cuspidata cuttings, having received twelve weeks of cold at five temperatures respectively (see page 20), were divided into two groups. One group was placed in the green- house under the natural daylength from mid-February until mid- April. During this period of time, natural daylength increased from 10 hours to 13% hours. The second group received supplemental light of approximately 200 ft. candles at dusk to insure a total of 15 hours daylength. Both groups of plants were maintained at a mini- mum day-night temperature of 60° F. :t 3° F. After eight weeks the percent buds broken and length of new shoot growth was measured. 22 Differential Chilling of Root and Shoot Preliminary studies indicate that the cold requirement for shoot growth is three weeks of cold at 35° F. Root activity is markedly stimulated after seven weeks of cold at 35° F. To separate these two responses, 12 one-year-old and 12 two-year-old cuttings of Taxus cuspidata were subjected to cold shoot and warm soil temperatures. An identical number of plants were subjected to warm shoot and cold soil temperatures. These plants had been grown under greenhouse conditions during the fall and early winter months prior to this treatment in February 1969. Twenty-four plants in group one (cold shoot, warm roots) were placed in a growth chamber maintained at 32° to 35° F. air temperature during the night and 35° to 39° F. day air temperature. In order to maintain warm soil temperatures, flat-sided rubber tubing encircled each pot and warm water from a controlled tem- perature water bath circulated through the tubing. Shredded styra- foam provided adequate insulation around the pot and soil surface. Soil temperatures were measured by thermocouples placed at various positions in the pot and at the soil surface—stem interface. Night soil temperatures ranged from 55° to 64° F. ; day soil tem- peratures ranged from 66° to 72° F. Light was supplied by eight fluorescent tubes at approximately 850 ft. candles for 9 hours the 23 first four weeks, 11 hours the following three weeks, and 13 hours for the remaining two weeks, corresponding to natural photoperiods. Twenty—four similar Taxus cuspidata cuttings were placed in the growth chamber next to the treated cuttings as cold shoot, cold root controls. The second group of twenty-four plants (warm shoot, cold roots) was placed in a root chamber which was mechanically refrigerated. These cuttings were set on a wire mesh screen in a refrigerated root chamber with the shoots projecting through a styrafoam lid. Shredded styrafoam insulated the soil and allowed the shoots to project into the greenhouse environment. Air day temperature in the greenhouse was maintained at a minimum 60° F. Soil temperature, as recorded by thermocouples at various soil depths, was maintained at 33° to 39° F. Natural photoperiods for the nine week duration of this study extended from 9 hours to 13 hours. Twenty—four similar Taxus cuspidata cuttings were placed adjacent to the root chambers to act as warm shoot, warm root controls. After nine weeks in their respective treatment chambers, all cuttings were removed and placed on greenhouse benches main— tained at a 60° F. minimum day-night temperature. Two weeks later, root activity was measured (see page 20). RESULTS AND DISCUSSION Root-Regenerating Potential Three-year—old Taxus hunnewelliana cuttings, lifted at biweekly intervals and root pruned, displayed a distinct periodicity in their root-regenerating potential. The number of new visible root initials produced was variable within each lifting date; however, a definite trend was evident (Figure 1). Considering all new root growth of any visible length which occurred during the six-week period in the growth chamber, perio- dicity in the root-regenerating potential was as follows: During the summer months (June 21 to August 30) an average of only 43 active initials per plant were produced (Table 1). As an indication of variability within lifting dates, seventy percent of the plants lifted during this period produced less than 20 visible active roots per plant, while the remaining thirty percent produced an average of 130 active new roots. The root-regenerating potential increased in the fall (September 13 to October 14) tO an average value of 85. There were continued increases in the root—regenerating potential 24 25 Figure 1. -- The yearly cycle of the root-regenerating potential of Taxus hunnewelliana. 2 3 Number of new root initials 2. 6756 + 28. 9646T - 1. 3270T + O. 0154T T = Time in weeks Sxy = 59. 4 R2 = .509** 26 TEMPERATURE- DCCREOEC F 0 o 0 . o o .8. 2 3?. °2 s a 3 a 3 122 f I I I I I I I I I I ’ 0’ O, G l ,l I. I / . -I I 0 ' I I ( - I i -\ . | . “3°C g l i . I I. II | . o d II C. I- ‘ \ \ ’- a ‘ ‘ ° 0 d E. .".' 2 \ \ - i .1 o \ . \ d 5 a E \ \ .. 2 Q g \\ °\ \ u: )- 5 5 \ 0 .Q g x 8 \ \ \ " " 5 '2' \I .\ °° §§ 3 \ \, ‘, I ‘- ° ' ° \ ' u ‘ g g g ‘\ \ 24 < < \ . . .4 I «'- I \ \. ° I ! i \\ \. \. d I ' \ ° I . 1 \ ‘ I I I \s\ ‘.\ -l \\ \ \.. ‘ \\ \ \ _I 1. 1 J I, | .I ° ' ,I .l d I? I d If] . I/ . I.) o/ I / /° . I - , I . ° 7 I l L A 1 J L 1 i l _1 L 8 2 8 2 a 3 2 8 3 8 ~ _ _ _ - - S‘WILINI 100' ‘3" IO UIIION SEP. 27 SEP. I! we.» (mus we. 2 Jams JULY 9 JUNE an «out 7 our 24 um 10 apn.zc APR. I: “0.29 mm: mum no.“ no. 2 «a.» ma. 9 01cm: o:c.4 NOV.25 NOV." 0:21.20 ocm4 LIFTIIO DATE 27 Table 1. -- The yearly cycle of the root-regenerating potential of Taxus hunnewelli ana. (Total number of new root initials produced 6 weeks after lifting. ) Number of New Initials Per Plant Lifting Date 1 2 3 4 5 Oct. 14 120 60 110 100 35 Oct. 28 0* 80 100 100 65 Nov. 11 121 22 24 86 53 Nov. 25 235 275 200 200 230 Dec. 9 230 210 300 100 200 Dec. 23 175 150 180 140 190 Jan. 5 185 106 70 12* 35 Jan. 19 50 295 0* 114 18* Feb. 2 115 85 76* _ 230 215 Feb. 16 280 317 293 192 409* Mar. 1 190 124 140 180 130 Mar. 15 43 209 95 131 85 Mar. 29 239 76 126 167 140 Apr. 2 265 155 425* 215 195 Apr. 26 33 5* 52 88 151 May 10 139 67 69 74 118 May 24 36 51 30 26 27 Jun. 7 1* 36 15 20 77 Jun. 21 158* 19 11 10 5 Jul. 5 2 7 12 4 7 Jul. 19 5 3 1 0 0 Aug. 2 0 5 24 0 12 Aug. 16 220* 115 280* 35 200* Aug. 30 98 32 0 5 8 Sep. 13 68 150 115 120 10 Sep. 27 65 95 126 61 26 *Residuals eliminated from regression analysis. 28 in late fall and early winter (October 14, November 11 and Decem- ber 9). A peak period in the regeneration Of new roots occurred during mid-winter (December 9 and February 16). An average of 183 new root initials per plant were produced during this peak period. This was followed by a sharp, prolonged drop in the root- regenerating potential between March 1 and June 7. In general this behavior closely corresponds to that reported earlier for Douglas- fir (Pseudotsufi menziesi, 46) and ponderosa pine (Pinus ponderosa, 43). Plants lifted between October 14 and early April were dormant or non—growing at the time of lifting. After six weeks in the growth chamber, less than 20% of the plants lifted between October 14 and December 23 produced sporadic new shoot growth. Normal bud expansion and shoot growth was produced on all plants in the growth chamber which were lifted on January 5 through early April. Terminal and lateral buds had expanded and new shoot growth had begun on all plants prior to lifting in mid-April. It is tempting in this case to imply an inverse relationship between bud expansion and root activity. Bud break in the growth chamber coincided with peak root-regenerating activity and precipi- tated a gradual decrease in the RRP from January 5 to late February. Thereafter, a more pronounced decrease in RRP occurred, reaching 29 a low in mid-July. It is generally acknowledged that in many woody plants there is a decrease in root activity in the spring with the onset of bud expansion. Plants lifted in the spring, in this case on April 12 when shoot growth was occurring, produced consistently fewer root initials from the time of bud break until mid-summer. The summer low in RRP cannot be explained by an inverse relation- ship between root and shoot growth. Soil conditions, temperature, and moisture content before lifting and in the growth chamber were favorable for root activity and were uniform throughout the experi- ment. Anatomy Dormant roots of Taxus cuspidata can be easily distinguished by the presence of a dark red root tip. This dormant structure is characteristic of all classes of roots: large primary roots (Fig- ure 2), short lateral roots, and roots just emerging from the cortex (Figure 3). Actively growing root tips lack pigmentation and appear a translucent white (Figures 2, 4). When a dormant root resumes activity, the old root cap cells are sloughed off and the active apex elongates. This may occur relatively quickly with the white active apex emerging, ringed by the dark red pigment of the old root cap cells which are rubbed off as the root elongates through the soil. More commonly, there is a gradual transition from dormant apex 30 Figure 2. -- From left to right: Dormant root tip, intermediate root tip, active root tip of Taxus cuspidata. Figure 3. —- Dormant lateral root tips of Taxus cuspidata. 31 32 Figure 4. -- Longitudinal section of an active root tip of Taxus cuspidata. A. Root Cap B. Endodermis C. Apical Initials Figure 5. -- Longitudinal section of a dormant root tip of Taxus cuspidata. A. Root Cap B. Endodermis C. Apical Initials D. Metacutized Cortical Cells 33 34 to active apex, resulting in an intermediate, non—growing condition (Figure 2). In this case, the red root cap cells are sloughed off before the active apex emerges and elongates. Frequently, these red root cap cells can be rubbed off on a non-dormant root which has not yet elongated. However, in a true dormant state, the root cap cells remain attached, protecting the dormant apex beneath. The internal structure of dormant roots offing cuspidata (Figure 5) is very similar to that described for noble fir (Abies procera) (50) and red pine (Pinus resinosa) (51). The endodermis (Figure 5), stained positively for suberin and lignin, extends distally to the area of the immediate apical derivatives. This metacutized layer continues downward in the root cap cells until the apical initials and their immediate derivatives are sur- rounded. The coloration noted at the tip of the dormant roots of Taxus cuspidata may be due to (1) the degradation of those cells external to the metacutized layer and effectively closed off from the rest of the root, (2) as well as a deposition of red pigment, possibly an anthocyanin. The structural modifications of a dormant root, as described above, create a relatively rigid encasement to protect the enclosed root initials from a variety of stresses in the soil environment. 35 Cold Requirement Root Dormancy The number of weeks necessary to meet the cold require- ment for roots is dependent upon the temperature to which the cuttings are exposed (Figures 6, 7). One through six weeks of cold 0 exposure at all of the specified temperatures (28° , 32 , 35° , 40° and 48° F. ) was not sufficient to break root dormancy and the mean per- cent active roots per plant for this six-week period was less than 0.50%. Root activity markedly increased after seven weeks of cold due primarily to the root response at 35° F. After nine weeks of cold at 28° , 32° and 40° F. , a significant increase in root activity occurred, more than doubling at 32° and 40° F. It was not until exposure to 12 weeks of cold at 48° F. that a significant response in root activity was produced. The final level of root activity attained after exposure to twelve weeks at 48° F. was significantly smaller 0 than the root activity produced after twelve weeks at 28° , 32 , 35° and 40° F. What level of root activity is necessary to consider root dormancy broken? A preliminary experiment was conducted the previous year (late February to late April, 1968) with one-year-old cuttings of Taxus cuspidata and root observations were made but no data was collected. 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Bennoomnoafi mo muoon o>EOo Eoonog zoo: I. .m. onswE 38 explosion of root activity occurred. Very little root activity was noted on plants chilled less than seven weeks. The response of one-year-old cuttings in the current experiment exposed to seven weeks of cold beginning in early December 1968 was less dramatic than was observed in the preliminary study (beginning in late Feb- ruary), although significant differences did exist between plants chilled for seven weeks and plants chilled less than seven weeks. The magnitude of response may be limited by available photosynthate. This point is clearly illustrated with two-year-old Bug cuspidata cuttings which were chilled beginning in late February until late April. Plants were chilled at four temperatures, 32° , 35° , 40° and 48° F. , and results compared ‘with non-chilled control plants which remained in the greenhouse at a 55° F. minimum night temperature (Figure 7). In general, root activity was higher for all weeks and all temperatures than was observed for the one-year-old cuttings. There was a gradual increase in root activity up to four weeks of chilling at 32° , 35° , 400 and 48° F. , followed by a decline in root activity after five weeks of chilling. After seven weeks of chilling at 32° , 35° and 40° F. , a highly significant increase in root activity occurred averaging 69% active roots. A gradual increase in root activity was observed at 48° F. , reaching maximum activity after eight weeks. The plants which remained in the greenhouse .‘t_r_.——2—1.tan. ’, . " . I J 39 exposed to a normal daylength regime cannot be considered a valid control comparison since they received normal light and chilled plants were in the dark for the duration of the chilling period. Root activity of unchilled plants was variable for the eight week duration of the experiment but never reached the maximum values obtained for plants chilled seven weeks at the three most responsive tem- peratures. It appears from this data that root activity is stimulated by cold treatment. The cold treatment for roots of Taxus cuspidata must be defined in terms of two parameters, time and temperature, and the resultant magnitude of response is dependent upon the age and vigor of the cutting. Shoot Dormancy The cold requirement for shoot dormancy of Taxus cuspi- data is a function of both time and temperature (Figure 8). Two weeks of chilling at 28° , 35° , 40° and 48° F. is not sufficient to break bud dormancy (Figure 10) and stimulate shoot growth (Fig- ure 9). Two weeks of chilling at 32° F. produces a significant increase in bud break; however, the greatest increase in percent buds broken occurs after three weeks of chilling. Three weeks of chilling at 28° , 35° , 40° and 48° F. increases the mean percent bud break by 40%. The mean percent buds broken is closely related to I if- 40 Figure 8. -- One-year-old Taxus cuspidata cuttings exposed to 1 through 4 weeks of chilling at 35° F. , as compared to cuttings which remained in the greenhouse (minimum temperature 65° F. ). 41 a cmm o a 08 3 5358 was; 42 .38: 58H 2.3 «m Bogota haucmouawwm mum on: nonmmp m .3 vmuwnmamm @8989: mmoFHI l I gown. woufi x “mmugmquEoH Coma/5.8m 3.8 .v 3&8 .m *3. .m ,3: .m 80:83.2:me no.“ cocoon “35:8th S .H mo .H N ”mxomg 8525mm **om.r **wpm.m *3. .m .453 A mocwowficmwm no.“ vmpom: 00:23me H Ho .8 mo .9 ”mcofimcgmmno 8:33ch Coma/pom **mo.pH **wbm.m *mm.mH *omm.H mocwoaficmfim no.“ “Boom: 828.8th Amcoflwcfimmno Lao.“ ”:3me ~03anva 8.8 8.8 3.3. 8.3. 8.8 8.8 3.2. 2.8 8.8. 8.2 85 25 M 8.8 8.8 8.8 8.8 8.2. 8.8 8.8 2.8 wwwflmmfi 8.8 85 85 .3. 8.8 8.8 8.8 8.2. 8.8 8.3. 8.3. 8.8 8.8 8.8.8.8 85 85 .8 8.8 2.8 2.8 8.8 8.8 8.8 8.8 8.2. 2.8 8.8"85 85 85 .8 8.8 8.8 8.8 8.8 8.8 8.3. 8.8 8.8 8.8 2.8 34mm 8.8 85 .8 8.8 8.8 8.8 8.8 8.3. 8.8 8.8 8.8 8.8 8.8 8.8“85 85 .8 M 2 2 S m w s 8 m. 5. m N a .m 88me acme—580.5. mo mxmm? mpsumpoagmh #:888on 9:58me -83 .858 mxooB 83835. .mmcfiuso 8.8.3.050 mnxwh. Eoimom$so Ho 788 fiaoum “002w mo 5mg: 552 u- .m opsmwh Am>8A 08m 85 8 88.8%? .nAucmoAAAchm 8.8 85A A8338 m ‘3 vmumuamm P8355: mmofilll “aim A. “31.30 .m *3 .m *m: .m meadouEmAm .8A A8888: 80.8.8AAAQ Ac .9 no.8 x "mmugmhmasmh A8838m Ac .8 no HA. M ”my—M83 .8838m 3.: .NA .3835 *AN .m *0: .A mocwouEmAm new A888: 828.8AAAQ Ac..H. no .H. ”mcoAAmPHmmnO 85339: A8838m :3 SN 38$ .m .33 .om .53 .A moQMQAAAcmAm new oovmmc 8288889 Amcoflmzmmno .38 5.8886 8.3:»:qu 3 4 ovdw owdm mmAm omhw oA.mw omda omém 3.3. ow.$ mmém ovAA om.m M Ayn .3. oo .3 mu .8 mm .3 mu .3 on .5 oo .wm on Am mmém co .3. mp .wm n co .m oo .o 03. :23 2.5. coda £23 omdm Slaw mmdm Show cog? 3.6m mm.NA._om.m mud 03. :22. coda mrAm 3.3 ooAm omém om.5 coda 3:2. ooAA. omfim _om.m cod 0mm 3.3. .363 mbdm npAm 3.5. 322. omdm mm.A.m mmdp code ondw “ macaw cod own 3. .mm mmdb mp .om mmdm om.mw oo .3. mm .mm mm .mm on .ow 3.2. on .mm _ mp .NA mad owm _ M NA AA 3 a w A. w m A. m N A .nA mmmgmwfl opsampwafimk ucwfiammuh. Ho 388.3 .888888 mugmhmnfimu .888 9883 838.5 .mwcAfi—AS «83.8.50 msxmfi Eofimomumco .8 “88.5 mung. 880me 582 .. .3 8.59m 44 subsequent length of new shoot growth. After three weeks of cold at 28° and 32° F. , length of new shoot growth increases twenty-fold and twelve-fold respectively. After four weeks of cold at 35° and 40° F. , the mean length of new shoot growth and percent buds broken are comparable to the response of plants chilled for three weeks at 28° and 320 F. A more gradual increase in shoot growth occurs after four weeks of chilling at 48° F. Shoot growth continues to increase through seven weeks of chilling at 28° and 35° F. , and then decreases significantly after eight weeks of chilling. A less abrupt decrease in shoot growth also is observed at 32° and 40° F. after eight weeks of chilling and at 48° F. after nine weeks of chilling. This decrease in shoot growth is accompanied by a concomitant increase in root activity. Length of new shoot growth was measured twelve weeks after each chilling treatment. This length of time was necessary to allow for a maximum response to the treatment. Since the twelve week growth periods did not occur simultaneously, shoot growth would vary with respect to photoperiod, light intensity, and maxi- mum air temperature. Daylength for this twelve week period in the greenhouse (early March to late May) increased from 10 hours to 14 hours. 45 ”mm 1].; ' . “a. x L . Daylength after Chilling Exposure of dormant Taxus cuspidata cuttings to twelve O 0 weeks of cold at the five specified temperatures, 28° , 32 , 35 , 40° and 48° F. , is sufficient to meet the cold requirement and to promote active shoot and root growth (Figures 6, 9). These tem— perature treatments exert a highly significant effect on the subse- quent length of new shoot growth and the percent of buds broken. However, the daylength which these cuttings are exposed to after cold treatment has no significant effect on subsequent shoot growth (Figure 11, Table 2) or bud break (Figure 12, Table 2). This study examines daylength effects on Taxus cuspidata after sufficient chilling to meet the cold requirement. Snyder (40) has shown that bud break did not occur on hardwood cuttings of Taxus cuspidata taken in mid-December and exposed to short photo- periods (8 hours). Similar cuttings under long photoperiods (18 hours) showed considerable shoot growth. The amount of chilling these plants had received in the natural environment before the cut- tings were taken may not have been sufficient to meet the chilling requirement. When plants have received insufficient or no chilling, long days may break bud dormancy (40). After the chilling require- ment for shoot growth has been met, daylength exerts no significant effect on bud break and length of new shoot growth. 46 Figure 11. -- Mean length of shoot growth (cm. , six replications) of one-year-old Taxus cuspidata cuttings exposed to 2 daylengths after 12 weeks of chilling at 5 temperatures. MEAN LENGTH SHOOT GROWTH (cm) 50" 45- 40b 35f 30' 47 25*- , SHORT DAY ’I —————— LONG DAY 20" L a or n I l 4 1 28’ 32‘ 35‘ 40‘ 48' TEMPERATURE (F) FOR l2 WEEKS 48 Figure 12. -- Mean percent buds broken (six replications) of one-year- old Taxus cuspidata cuttings exposed to 2 daylengths after 12 weeks of chilling at 5 temperatures. BUDS BROKEN MEAN PERCENT IOO - 95'- 90b 85" 49 28‘ SHORT DAY ------ LONG DAY 1 l 1 32‘ 35‘ 40‘ TEMPERATURE (F) FOR l2 WEEKS 48’ -‘J fi‘gnlfi" ~ . . 50 Table 2. -- Effect of daylength on shoot growth of Taxus cuspidata after 12 weeks exposure to 5 temperatures. 1W“ I“); _ Short Day Long Day Tenux 33 38.7 75 25.6 39 31.7 78 27.6 280 100 40.5 67 34.1 28 7.6 73 32.9 100 36.0 57 14.2 57 20.3 100 19.1 39 40.0 89 54.8 100 50.2 85 53.9 32. 100 35.9 100 35-4 100 65.8 100 46.0 100 54.2 100 45.6 100 32.8 73 37.3 32 51.4 90 47.7 92 55.3 100 36.4 35. 100 36.1 88 41-7 90 51.3 100 27.9 35 52.4 100 44.0 100 41.0 100 44.6 100 53.1 33 36.3 89 46.1 39 43.4 40° 33 53.0 100 58.3 100 35.0 80 53.9 37 43.0 100 39.7 100 51.6 90 61.4 91 52.2 100 53.0 100 47.1 100 63.7 48° 87 66.5 33 43.8 73 43.4 93 54.0 92 57.9 100 33.7 100 33.0 89 50.8 51 1 Differential Chilling of Shoot and Root The cold requirement for root dormancy of Taxus cuspidata (Figure 6) had been established as seven weeks of chilling at 35° F. Shoot dormancy is broken after three weeks of chilling at 32° - 35° F. (Figure 9). This experiment was designed to determine whether the shoot or root "sensed" the chilling treatment for root dormancy. The nine week chilling period in this experiment was sufficient to meet the cold requirement for both shoots and roots. These two shoot and root responses are separated into four treatments as follows: Cold Shoot Warm Roots Treatment 1: Warm Shoot Treatment 2. Cold Roots Cold Shoot Treatment 3. Cold Roots Warm Shoot Treatment 4: Warm Roots Cold Shoot . . . . . . Treatment 1 (Warm Roots) IS highly Sigmficantly different from the other three treatments in either percent dormant or percent . . Warm Shoot active roots. Comparisons among treatment 2 ( Cold Roots)’ treat- Cold Shoot Warm Shoot ment 3 (Cold Roots) and treatment 4 (Warm Roots) show that all are highly significantly different at the 1% level except differences 52 “PISL 7‘0- Table 3. —- Differential chilling of shoot and root. (Plants chilled for 9 weeks. ) Treatment Percent Percent Active Roots Dormant Roots 1' v53; 31:13:63 ”~35 61.15 2' “£33833? 63- 62 36. 38 3- 5313 $335; 75- 33 24. 67 4' 3):: 312:2; 16- 79 83.21 Difference Needed for Significance hsd.05 lvs 2, 3or4 11.84 11.84 hsd . 01 1 vs 2, 3 or 4 14.54 14.54 hsd .05 among 2, 3 and 4 9. 92 9. 92 hsd .01 among 2, 3 and 4 12.19 12.19 53 between treatments 2 and 3, which are significant at the 5% level (Table 3). Ranked in order of greatest percent root activity to least percent root activity, the treatments are as follows: Cold Shoot _ Warm Shoot _ Cold Shoot _ Warm Shoot Cold Roots Cold Roots Warm Roots Warm Roots This would seem to furnish good evidence that cold treat- ment to shoots increases root development but does not substitute directly for the chilling requirement of the roots themselves. Cold Shoot Warm Roots Comparisons between treatment 1 < ) and treat- ments 2, 3 and 4 are based on unequal observations. Eleven of twenty-four plants in treatment 1 were not included in the root activity comparisons because the root system appeared dead and/ or the shoot had browned—off and buds were not viable. This occurred only with treatment 1, and plants in the same growth Cold Shoot Cold Roots Comparisons among the other three treatments (2, 3 and 4) included chamber (treatment 3, ) were not adversely affected. the total of twenty-four observations per treatment; no adverse effects were noted. Due to the unequal observations in treatment 1, statistical comparisons between age of cuttings were not made. The table presented below, however, indicates that little difference exists between one-year—old rooted cuttings and two-year-old rooted cuttings for all treatments. 54 Percent Active Roots Treatment Total One-Year-Old Two-Year-Old Cuttings Cuttings v53; 81:12:; 38. 85 36. 60 41.00 122313;: 63. 62 71.16 56. 08 8:1: :23: 75. 33 74. 33 76. 33 warm Show 16.79 17.58 16.00 Warm Roots CONC LUSIONS Root dormancy is a natural phenomenon; it does exist in certain species of M. The anatomical study discussed in this paper clearly shows that a distinct structural modification exists in dormant roots. Such a modification may, in fact, be a protective adaptation to unfavorable growing conditions, but is not necessarily brought about by unfavorable conditions. It has been proven in other work with pine seedlings (13, 23) that a metacutized, non-growing root tip is formed due to water stress. Favorable moisture and temperature conditions for root growth were maintained in this study and yet non-growing, dormant root tips were formed. Apparently, environmental stress is not a prerequisite for formation of dormant root tips in Taxus cuspidata or certain species of Pinus (13, 23). In breaking dormancy, the winter environment in the tem- perate zone chills the above and below ground portions of the plant, initiating internal changes in the chilled apices (34). Plants in this post—dormant phase resume growth with favorable conditions in the spring. Dormant buds and dormant roots Of Taxus cuspidata are responsive to chilling. Two separate and distinct chilling 55 56 requirements exist for bud dormancy and root dormancy. Three weeks of chilling at 280 and 32° F. are required to break bud dor- mancy and at least seven weeks of chilling at 35° F. are necessary to break root dormancy. Dormant buds are responsive to lower temperatures than dormant roots. This may be an important adap- tive feature of dormant buds as air temperatures fluctuate much more than soil temperatures throughout the winter (Figure 1). A soil mass conducts heat more slowly and therefore cools and warms at a slower rate than the surrounding atmosphere. Soil temperatures lag slightly behind warming air temperatures in the spring and like- wise, in the fall and earlywinter, soil temperature gradually decreases, cooling more slowly than the surrounding air tempera- ture. Root and shoot dormancy in Taxus cuspidata can be viewed as two separate phenomenon operated by two different systems responsive to different periods of chilling at different temperatures. The cold requirement for the roots appears to be "sensed" by the roots themselves; however, the presence of a physiologically non- dormant bud significantly increases root activity (Table 3). Further effects of the shoot on root activity are illustrated by the increase in root activity when only the buds are chilled (Table 3). These two separate systems, one controlling root activity and one controlling 57 shoot activity, are evidently woven together or linked in some manner, as shown by the effect of one upon the other. Chilling may not be the only factor involved in breaking root and shoot dormancy. Photoperiod may break bud dormancy in Taxus cusglata (40) and consequently have an effect on root activity before the plants are chilled. The results of the study with two-year-old Taxus cuspidata cuttings (Figure 7) may indicate a unique interaction of the effect of photoperiod and chilling on root activity. The data are difficult to interpret on the basis of chilling alone. There appears to be an additive effect of increasing daylength and chilling on root activity after four weeks. The decrease in root activity after five weeks of chilling may be due to a competitive effect of active shoot growth. The greatest increase in root activity occurs after seven weeks of chilling when the cold requirement for root growth has been fulfilled. Root and shoot dormancy are related, at least in part, to the root-regenerating potential (RRP). Plants lifted in the late fall and early winter have attained a high RRP. Low winter soil tem- peratures breaking root dormancy may have an additive effect on the RRP. The cold requirement for bud break has been completed by January 5 (as evidenced by bud break in the growth chamber). With the increase in shoot growth after bud dormancy has been broken, there is a concomitant decrease in the RRP. Bud break 58 and shoot elongation occurred in the natural environment by April 12. The decrease in root activity following bud break, both in the growth chamber and in the natural environment, may be due to increased competition with shoot growth for available carbohydrates and other necessary growth factors. Shoot elongation in Taxus hunnewelliana is generally completed by late June. Therefore the summer low in the RRP is not related to competition with active shoot growth. The early rise in the RRP from August 30 to the middle of November cannot be attributed to the breaking of root dormancy as soil tem- peratures are as yet too warm to be effective in meeting the cold requirement. The summer low in the RRP and the early fall rise in the RRP may be regulated by an environmentally triggered internal system which affects root initiation or the elongation of root initials which were present for several growing seasons. Daylength may be the controlling factor regulating the RRP during this period. In July and August the photoperiod decreases from 15 hours to 13% hours when the RRP is at its lowest. As daylength decreases from 12 hours in September to 9% hours in early November, the RRP in- creases sharply. The critical daylength affecting this increase in RRP may be approximately 10 hours. It would be possible to test this hypothesis by measuring the RRP of plants maintained under long—day (15 hours) conditions from July through November. 59 The relationship of root dormancy to root initiation is unclear. It has not been determined when new root initials are formed. The new roots regenerated by the severely pruned root system may have been previously formed in the pericycle and remained imbedded in the cortex of the root, the severe pruning having stimulated the growth of these imbedded new roots. Or, root initials once formed, may elongate immediately and be exposed to the soil environment. If the latter proves to be correct, the RRP is a measure of root initiation at the time it occurs in the pericycle. Root initials formed in the pericycle may become dormant before breaking through the cortex of the root. This seems very likely as red dormant root tips were observed on large white roots to occur flush with the root surface, not protruding through the outer layer of root tissue. If the regenerated root system is produced from newly formed root initials, plants may be dug bare-root during a high RRP period and placed in cold storage until transplanted in the spring. The careful protection of the finer roots may not be neces- sary. If root initials are preformed in prior seasons, it would be necessary to protect the finer roots of lifted stock more carefully. Otherwise, the removal of the finer lateral roots would severely limit the regenerative capacity of the remaining root system. Exact optional dates for lifting are difficult to predict from one year to the next due to changing weather conditions. If, however, 60 further research substantiated the proposed daylength mechanism for the increase in RRP, exact lifting dates could be predicted. Plants lifted during the ”optimal" daylength period could be placed in cold storage to fulfill the shoot and root cold requirement resulting in maximum transplanting survival. LITERATURE CITED 10. LITERATURE CITED Chandler, K., M. H. Philp, W. P. Tufts, and G. P. Weldon. 1937. Chilling requirements for opening of buds on deciduous orchard trees and some other plants in California. Calif. Agr. Expt. Sta. Bul. 611, 63 pp. Corville, F. V. 1920. The influence of cold in stimulating the growth of plants. Jour. Agr. Res. 20:151-160. Crider, F. J. 1928. Winter root growth of plants. Science. 68:403-404. Downs, R. J. Photocontrol of growth and dormancy in woody plants. In: Tree Growth, T. T. Kozlowski, ed. Ronald Press C0,, New York. pp. 133-148. 1962. . Eliasson, L. 1968. Dependence of root growth on photosynthesis in Populus tremula. Physiologia Plantarum. 21:806-810. Farmer, R. E. 1968. Sweetgum dormancy release: Effects of chilling, photoperiod, and genotype. Physiologia Plantarum. 21(6):1241-1248. Gustafson, F. G. 1938. Influence of the length of day on the dormancy of tree seedlings. Plant Physiol. 13:655-658. Harris, G. H. 1926. The activity of apple and filbert roots especially during the winter months. Proc. Amer. Soc. Hort. Sci. 23:414-422. . Head, G. C. 1966. Estimating seasonal changes in the quantity of white unsuberized root on fruit trees. J. Hort. Sci. 41:197- 206. Howard, W. L. 1910. An experimental study of the rest period in plants. The winter rest. Mo. Agr. Expt. Sta. Res. Bul. 1, 105 pp. 61 11. 12. 13. 14. 15. 16. 17. 18. 19. 20. 21. 22. 62 Jensen, W. A. Botanical Histochemistry. W. H. Freeman Co., San Francisco. 325 pp. 1962. Kaufmann, M. R. 1968. Water relations of pine seedlings in relation to root and shoot growth. Plant Physiol. 43(2):281-288. Kramer, P. J. , and H. C. Bullock. 1966. Seasonal variations in the proportions of suberized and unsuberized roots of trees in relation to the absorption of water. Amer. Jour. Bot. 53(2): 200-204. Kramer, P. J., and T. T. Kozlowski. Physioloégy of Trees. McGraw-Hill Book Co. , New York. 642 pp. 1960. Krugman, S. L., E. C. Stone, and R. V. Bega. 1965. The effects of soil fumigation and lifting date on the root-regenerating potential of Monterey pine planting stock. J our. Forestry. 63: 114-119. Krugman, S. L., and E. C. Stone. 1966. The effect of cold nights on the root-regenerating potential of ponderosa pine seed- lings. Forest Science. 12(4):451-459. Lamb, R. C. 1948. Effect of temperatures above and below freezing on the breaking of rest in the Latham raspberry. Proc. Amer. Soc. Hort. Sci. 51:313-318. LeClerg, E. L., W. H. Leonard, and A. G. Clark. Field Plot Technique. Burgess Publ. Co., Minneapolis, Minn. 373 pp. 1962. Luckwill, L. C. 1960. The physiological relationship of root and shoot. Hort. Sci. 14:22-26. Lyford, W- H. , and B. Wilson. 1966. Controlled growth of forest tree roots: technique and application. Harvard Forest Paper No. 16. McDougal, W. B. 1916. The growth of forest tree roots. Amer. Jour. Bot. 3:384-392. Merritt, Clair. 1968. Effect of environment and heredity on the root growth pattern of red pine. Ecology. 49(1):34-40. 23. 24. 25. 26. 27. 28. 29. 30. 31. 32. 33. 63 Meyer, M. M., and H. B. Tukey, Jr. 1967. Influence of root temperature and nutrient applications on root growth and mineral nutrient content of Taxus and Forsythia plants during the dor- mant season. Proc. Amer. Soc. Hort. Sci. 90:440-446. Mullin, R. E. 1967. Root exposure of white spruce nursery stock. Forestry Chronicle. 43(2):155-—160. Nienstaedt, H. 1966. Dormancy and dormancy release in white spruce. Forest Science. 12:374-384. Nienstaedt, H. 1967. Chilling requirement in seven Picea species. Silvae Genet. 16:65-68. Nightingale, G. T. 1935. Effects of temperature on growth, anatomy, and metabolism of apple and peach roots. Bot. Gaz. 96:581-639. ' Richardson, S. D. 1956. Studies of root growth in Acer saccharinum L III. The influence of seedling age on the short- term relation between photosynthesis and root growth. Proc. Ned. Akad. van Wet. C59(3):416-428., Richardson, S. D. 1956. Studies of root growth in Acer saccharinum L IV. The effect of differential shoot and root temperatures on root growth. Proc. Ned. Akad. van Wet. C59(3):428—438. Richardson, S. D. 1956. Studies of root growth in Acer saccharinum L V. The effect of a long-term limitation of photo- synthesis on root growth rate in first-year seedlings. Proc. Ned. Akad. van Wet. C59z694-701. Richardson, S. D. 1957. Studies of root growth in Acer saccharinum L VI. Further effects of the shoot system on root growth. Proc. Ned. Akad. van Wet. C60z624-629. Richardson, S. .D. 1958b. Bud dormancy and root development in Acer saccharinum L. In: The Physiology of Forest Trees. Ed. K. V. 'I'himann. Ronald Press Co., New York. pp. 409— 425. Romberger, J. A. Meristems, Growth, and Development in Woody Plants. U.S.D.A. Forest Service Tech. Bul. No. 1293. 214 pp. 1963. 34. 35. 36. 37. 38. 39. 40. 41. 42. 43. 44. 45. 64 Rogers, W. S. 1939. Root studies VIH. Apple root growth in relation to rootstock, soil, seasonal and climatic factors. Jour. Pomol. and Hort. Sci. 17:99-130. Rogers, W. S., and G. A. Booth. 1960. The roots of fruit trees. Hort. Sci. 14:27-34. Samish, R. M. 1954. Dormancy in woody plants. Ann. Rev. Plant Physiol. 5:183-204. Schubert, G. H., and F. J. Baron. 1965. Nursery tempera- ture as a factor in root elongation of ponderosa pine seedlings. Pacific Southwest Forest and Range Experiment Station. U. S. Forest Service Research Note PSW—66. Smith, H., and N. P. Kefford. 1964. The chemical regula- tion of the dormancy phases of bud development. Amer. J our. Bot. 51(9):1002-1—12. Snyder, W. 1955. Effect of photoperiod on cuttings of Taxus cuspidata while in the propagation bench and during the first growing season. Proc. Amer. Soc. Hort. Sci. 66:397-402. Steel, R. G., and J. H. Torrie. Principles and Procedures in Statistics. McGraw-Hill Book Co. , New York. 481 pp. 1960. Stone, E. C. 1955. Poor survival and the physiological con- dition of planting stock. Forest Science. 1(2):90—94. Stone, E. C., and G. H. Schubert. 1959a. Root-regeneration by ponderosa pine seedlings lifted at different times of the year. Forest Science. 5:322-332. Stone, E. C., and G. H. Schubert. 1959b. The physiological condition of ponderosa pine (Pinus ponderosa Laws.) planting stock as it affects survival after cold storage. Jour. Forestry. 57:837-841. Stone, E. C., and R. W. Benseler. 1962. Planting ponderosa pine in the California pine region. Jour. Forestry. 60(7):462- 466. Stone, E. C., J. L. Jenkinson and S. L. Krugman. 1962. Root- regenerating potential of Douglas-fir seedlings lifted at different times of the year. Forest Science. 8(3):288-297. 46. 47. 48. 49. 50. 51. 52. 53. 65 Stone, E. C. 1967. The root regenerating capacity of seedling transplants and the availability of soil moisture. Annals of Arid Zone. 6(1):42-57. Vegis, A. 1956. Formation of the resting condition in plants. Experientia. 12:94-99. Vegis, A. 1964. Dormancy in higher plants. Ann. Rev. Plant Physiol. 15:185-224. Wilcox, H. E. 1954. Primary organization of active and dor- mant roots of noble fir, Abies procera. Amer. Jour. Bot. 41(10):812-821. Wilcox, H. E. 1968. Morphological studies of the root of red pine, Pinus resinosa. I. Growth characteristics and patterns of branching. Amer. Jour. Bot. 55(2):247-254. Wilson, B. F. 1964. Structure and growth of woody roots of Acer rubrum L. Harvard Forest Paper No. 11. Worrall, J. , and Francois Mergen. 1967.» Environmental and genetic control of dormancy in Picea abies. Physiologia Plantarum. 20:733-745. Zaerr, J. B. 1967. Auxin and the root-regenerating potential in ponderosa pine seedlings. Forest Science. 13(3):258-264. APPENDIX TABLES 66 :52 .2 3:89:56 5:353. 8mm .3 u Tagasmm 36 32m 285% mmom .o n mmnnacmwoammoo “83.29500 53332 $30 .o can .7 36m .wm mag .m masowoflmoov month. Noam. 685. 3me:00 musmfioamoou sowmmmuwom mmmémmm www.mvmwmm m: .3ng .3325 .o moo .mm can .552 mam .333. m sowmmopmmm momdmmzm m: 130B 853meme oflmflmam Puma—Um mohmsum Eoummhh mosmflhw> mo monsom mumawxoumadw m 532 mo 83m mo mmmpmom coflmmonwom Zahm>0 .HoM oonEm> mo 3.6.3.344 new 3885600 28339300 @3338 .musowoummoo sowmmonmmu .Amsmwfimkogas .opmfifimo mo .8qu cumvsmum msxmh. mo 33:32“ wcfimnonommuuuoob cowmmmuwoh 3.295 .HoWooawflwur Mo 29393411 )2 3an 67 .26. OS .E-anwE hams...- Hmiz o... .38.??? mam .mmmmm 3N ESP 2.- .3 mm... .32: 9: Am: 82m .38 .o 25 .H mm. .2. 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