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'1.1I 1 1111111111111111111 1 IIII ...~IIII'III II. . 1 I1 ‘1'1111111111 1. .1111111'111111111t111111 11111111 "1 11:r-1;1:11.1~1 I 1..) 11.1 ”11‘:1 I 1 .1'I11II.' .1111 I111 1! 1 I ‘1 11.11 1 11.. 1 . .1 ~1"..‘! ‘ngg....1‘..11 _T.1 111 1 1L11I. 111111111111111111I1 11.1 I i. 111111 11111111 111111111111111111111 .1 Tussle This is to certify that the thesis entitled A Genetic Study of Salt Tolerance in Barley (Hordeum Vulgare L.) presented by Azzildeen M. Al—Shamma has been accepted towards fulfillment of the requirements for MOS. . degreem Crop & Soil Science Dag/2W]? ‘ WW I 0-7639 -__..__*-_—M—."fmr_..__—...__.— ._....—.-M..——._‘.—- ._ ____ OVERDUE FINES ARE 25¢ PER DAY PER ITEM Return to book drop to remove this checkout from your record. A GENETIC STUDY OF SALT TOLERANCE IN BARLEY (HORDEUM VULGARE L.) By Azzildeen M. Al-Shamma A THESIS Submitted to Michigan State University in partial fulfillment of the requirements for the degree of MASTER OF SCIENCE Department of Crop and Soil Science 1979 ABSTRACT A GENETIC STUDY OF SALT TOLERANCE IN BARLEY (HORDEUM VULGARE L.) by Azzildeen M. Al-Shamma Six varieties of barley of different origin were used to show the effect of salt on germination stage and mature- plant stage. A 6 X 6 diallel cross was made to study the genetic basis of salt tolerance during germination stage using Jinks—Hayman diallel cross analysis. Varietal differences for salt tolerance were obvious in both stages of growth. There was no correlation between germination stage and mature-plant stage for salt tolerance suggesting the possibility of the presence of at least two sets of genes. The genetic study, using F3 populations, indicated the presence of two different genetic systems for salt tolerance: in barley. One was found in California Mariout, in which salt tolerance is controlled by recessive genes. The second was found in the rest of the varieties in which the tolerance is controlled by dominant genes. To my parents ....... ACKNOWLEDGEMENTS The author wishes to express his sincere appreciation to Dr. J. E. Grafius for his guidance and encouragement throughout this study and for his constructive criticism in the preparation of this manuscript. Gratitude is also expressed to Dr. D. Smith and Dr. C. Cress for serving as Guidance Committee members and to Dr. C. M. Harrison for his review of the original manu- script. Appreciation is also extended to Dr. M. W. Adams for his critical evaluation of this study. The author greatly appreciates the financial support from the Government of Iraq for making this study possible. He is grateful to his mother and all members of his family for their moral support. Special thanks to his brother, Dr. A. R. Al-Shamma, for his support throughout this study. iii TABLE OF CONTENTS Page LIST OF TABLES . . . . . . . . . . . . . . . . . . . . v LIST OF FIGURES . . . . . . . . . . . . . . . . . . . vii INTRODUCTION . . . . . . . . . . . . . . . . . . . . . 1 REVIEW OF LITERATURE . . . . . . . . . . . . . . . . . 4 MATERIALS AND METHODS . . . . . . . . . . . . . . . . 16 RESULTS . . . . . . . . . . . . . . . . . . . . . . . 22 DISCUSSION . . . . . . . . . . . . . . . . . . . . . . 48 SUMMARY AND CONCLUSIONS . . . . . . . . . . . . . . . 55_ LITERATURE CITED . . . . . . . . . . . . . . . . . . . 58 iv LIST OF TABLES Table 10. Barley varieties that were used in this study . Mean squares of salt tolerance scores as represented by dry weights of the six varieties of barley at maturity . t values for the difference between two slopes for the six varieties of barley for dry weight, fresh weight, height and tiller number . . . . . . . . . . . . . . . . Mean squares of salt tolerance scores as represented by fresh weights of the six varieties of barley at maturity . Mean squares of salt tolerance scores as represented by heights of the six varieties of barley at maturity . Mean squares of salt tolerance scores as represented by tiller number of the six varieties of barley at maturity . Average moisture content (grams) per gram of dry matter of the six varieties of barley at three levels of salinity . . . Mean squares of salt tolerance scores as represented by germinating seeds of the six varieties of barley . t values for the difference between two slopes for the six varieties of barley for germination . The correlation among regression coeffi- cients for germination % and mature-plant characteristics . Page 16 22 26 29 32 35 36 37 41 42 Table Page 11. Mean squares for salt tolerance scores in the germination stage of the F3 of the 6 x 6 diallel cross set . . . . . . . . . . . 43 12. Average germination percentages of the F3 of the 6 x 6 diallel cross set . . . . . . . 45 vi Figure 1. Effect of NaCl concentration on the total dry weight of 6 barley varieties . 2. Effect of NaCl concentration on the dry weight of 6 barley varieties . 3. Effect of NaCl concentration on the total fresh weight of 6 barley varieties . 4. Effect of NaCl concentration on the fresh weight of 6 barley varieties . 5. Effect of NaCl concentration on the total height of 6 barley varieties . 6. Effect of NaCl concentration on the height of 6 barley varieties. . . . . 7. Effect of NaCl concentration on the total number of 6 barley varieties . 8. Effect of NaCl concentration on the tiller number of 6 barley varieties 9. Effect of NaCl concentration on the total germination percentage of 6 barley varieties . 10. Effect of NaCl concentration on the germina- tion percentage of 6 barley varieties ll. Wr/Vr graph analysis of germination percent- LIST OF FIGURES age of the F3 of the 6 x 6 diallel cross set . vii Page 24 25 27 28 3O 31 33 34 38 39 47 INTRODUCTION Soil salinity is common in some arid and semi—arid regions of the world where rainfall is insufficient to leach the salt out of the root region. Soils severely affected can easily be recognized by a thin layer of white powdery material covering the soil surface. Chloride, sulfate, carbonate and bicarbonate salts of sodium, calcium, magnesium and potassium are generally found in saline soils. The concentration of these salts varies from.one region to another. These salts have a high degree of solubility in water which makes them easily re- moved from rocks and soils by erosion. Due to this process, irrigation water may be high in salt content and become a major source of salts added to the soil. Problems develop from salts already in the soil, especially in arid and semi-arid regions where salts accumulate under certain environmental conditions. One is soil structure in which a layer of clay lies at various depths beneath the surface developing a poor drainage system in the soil. As a result, a salt solution would not pene- trate through this layer and salt accumulates. Another factor is a high ground water table which provides a poor drainage system. In either case, water will not penetrate this layer. Capillary movement might carry the salts to the soil surface or allow lateral movement. As the water reaches the soil surface, it evaporates, leaving the salts to accumulate near the plant root zone. High salt content in the soil reduces the availability of water to the plant as the osmotic pressure within the rooting medium increases and becomes higher than that in the plant tissues. It also affects the nutrient uptake by the plant. As a result, plant growth is reduced signifi- cantly causing a drastic reduction in yield. Farming this kind of land becomes nonprofitable and leads farmers to leave their land since reclamation and maintenance are very costly. With the increase in the population of the world and the demand for food, agricultural land has become too valuable to lose. Some farmers have become aware of this fact and started to grow salt tolerant species. According to studies in this field, salt tolerance was found to be a heritable characteristic in plants. Proper breeding programs then should be effective in devel- oping or improving some important economic crops. Extensive research is definitely needed to keep the salt concentration down to a level where a profitable farming system could be attained. To study the tolerance of plants to salt, all stages of growth should be considered since the reaction to salt might be different from one stage to another. This study investigated the characteristics of salt tolerance in barley plants (Hordeum vulgare L.) during germination and later stages of growth. REVIEW OF LITERATURE A saline soil is one with sufficient soluble salts to injure or reduce the growth of many plants. Irrigation water is a major factor in causing salinity problems. Depending on the geological structure of the soil through which streams and rivers pass, different water has differ- ent salt content. Allison (5) reports that most irrigation waters contain 0.1 to 5 tons of salt per acre-foot (70 to 3500 ppm). He classifies irrigation water on the basis of electrical conductivity measurements into four classes: Low salinity, medium salinity, high salinity, and very high salinity, the dividing points between classes being 250, 750, and 2250 umho/cm. This range includes water that can be used for irrigation of most crops on most soils to water harmful to use for irrigation under ordinary condi- tions. In a given soil, the cations calcium, magnesium, and sodium and the anions chloride, sulphate, bicarbonate, and carbonate are generally predominant (5, 17). Potassium occurs but in lesser proportions than any of the cations mentioned (5). Proportions of these ions may vary con- siderably among saline soils. Addition of fertilizers may cause an increase in the variety of ionic species present in excess (17, 39). The presence of these salts in the soil causes the osmotic pressure of the soil solution to rise high enough to create a water stress condition. The osmotic pressure within the root hairs of most plants is usually about 2 bars (40). In saline soils, the osmotic pressure may rise far beyond 2 bars, and unless plants adjust to the osmotic pressure within their root hairs, they would not survive. Berstein (13) provided evidence that the water adsorption capacity is relatively unaffected by salin- ity. He related the reduction in plant growth associated with osmotic stress to building up of the osmotic pressure of developing cells to meet the increasing osmotic pressure of the rooting medium and still maintain turgor. He then defines salt tolerance according to his theory as, "the degree to which osmotic adjustment can be made without sacrifice to growth". It was reported earlier, however, by Eaton (27) that as the salinity of the medium increases, the osmotic pressure of the leaves or aboveground parts of the plant increases. This results in the maintenance of essentially a constant gradient between medium and plant. Hayward and Spurr (34) measured the rate of entry of water into corn (Zea maize) roots under different osmotic pres- sures. A significant reduction in the rate of entry was found in both non-conditioned and pre-conditioned plants to high osmotic concentration of the substrate. Seeds of alfalfa (Medicago sativa) showed a reduction in hydration as the concentration of the substrate increased with either sodium chloride or mannitol (49). Efficiency of water use is also affected by salinity. The water requirements of wheat and saltbush decreased as the salinity increased, i.e., less water was used per gram of dry matter produced (26). In contrast, in a later study, Eaton (27) noticed an increase in water use for mixed culture of eight crops at higher levels of salinity than at moderate levels. This probably was due to accumulation of additional salts in plant tissue which might have led to the uptake of more water. The osmotic pressure concept in describing plant growth depression is predominant in the literature. Plants in different studies were exposed to different levels of artificial osmotic pressure using mannitol and some other salts. Isosmotic pressures induced by mannitol and salts seem to have different effects on plant growth. Uhvits (49) using Arizona grown Chilean alfalfa seed, observed that germination percentage was reduced much more in sodium chloride than in mannitol at equal osmotic pressures. Different salts may have different effects on plant growth. Gauch and Wadleigh (30) showed that sodium chloride, calcium chloride, and soidum sulfate at isosmotic concentrations had similar effects on the growth of red kidney bean plants (Phaseolus vulgaris L.), whereas with magnesium.chloride and magnesium sulfate, growth was depressed markedly. This brings out the effect of specific ions. Kofranek gt 31. (39), in greenhouse operations, noticed that heavy application of fertilizer developed a specific ion effect on the growth of Chrysanthemum (Chrysan— themum.morifolium) associated with high levels of ammonium or magnesium salts. In both studies, magnesium as the specific ion provided additional depression in plant growth. Higher concentrations of magnesium in the soil may cause toxicity to the plants (35). This effect, however, can be overcome if a moderately high concentration of calcium and potassium are present in the soil. Excessive concentrations of calcium, on the other hand, may depress the uptake of other cations. . Bernstein and Ayers (15) in studying the salt tolerance of five varieties of carrot (Daucus carota), noticed that at a given level of salinity using sodium chloride and calcium chloride, the sensitive varieties accumulated more calcium but less potassium. Hayward and Wadleigh (35), on the other hand, found that the presence of excessive sulfates decreased the uptake of calcium but promoted the uptake of sodium. As a result, sodium toxicity was induced (20). Fruit crops are found to be very sensitive to sodium. Sodium injury in almonds was found in non—saline soils containing less than 5 percent of exchangeable sodium (17). Toxicities, of sodium and chloride ions, however, were considered as major factors in salt damage to specifically sensitive fruit crops. Chloride is one of the major anions found in saline soils. It occurs in the form of salts with the cations sodium, calcium, and magnesium. It may be present in traces up to a few m.eq./l. in non-saline soils to about 100 m.eq./l. in saline soils (13, 14, 15). Under saline conditions, plants may accumulate up to 150 m.eq./100 gm or more chloride in their leaves. Allison (5) reports that chloride may accumulate to about 1 or 2 percent of the dry weight when the concentration in the root medium ranges from 700 to 1500 ppm. At those concentrations, plants show toxicity symptoms in which marginal burn of the leaves occurs, causing leaf drop, twig die-back and even death of the plant. Chloride was found to have little or no effect on the uptake of the essential anions phosphate, nitrate, and sulphate even at high concentrations (3000 ppm) in the rooting medium (45). Brown gt a1. (20) found that stone-fruit trees take up about twice as much Cl' per m.eq. of C1" in the nutrient solution from calcium chloride than from sodium chloride. It was reported earlier, however, that the uptake of chloride from added calcium and sodium chlorides was found to be equal for most plants studied. The mechanism of salt tolerance in the case of Na may be based on the particular plant or species to keep the proper Na level, in the leaf tissue below toxic levels and compensating for the lower water potentials associated with salinity by increasing levels of organic solutes in the tissue (46). Scholander, gt El- (47) and Atkinson, gt El- (6) have shown two different mechanisms of salt tolerance in taxonomically diverse mangroves. The first is concerned with excluding the salt in seawater by the roots: this was found in Rhizophora and Bruguriera. The second involves taking up the salt and excreting it by special glands on the leaves before toxic levels are reached in the shoot, examples: Aegiceras, Aicennia and Aegialitis. In a study involving six clones of Festuca rubra and four of Agrostis stolonifera, tolerance was associated with the restriction of Na+ and Cl’ accumulation in the shoots and the maintenance of almost constant concentrations in the roots over the salinity levels of 0, 25%, 50%, and 75% seawater (32). This might be in favor of the excluding theory since the tolerant festuca clones were considered as an effective excluder at both low and high salinities. Much of the research done on salt tolerance in plants has been during germination and early growth stages (1, 8, 12, 21, 22, 23, 24, 36, 44, 48). Soil salinity seems to have a significant effect on plant growth in general. It decreased the percentage of germinating seeds, increased the time of germination, and delayed the emergence of seedlings. Donovan (24) found that barley seeds required an additional 3-5 days to germinate in a saline culture. Maliwal and Paliwal (42) also stated that germination percentage of wheat (Triticum aestivum) and barley at different salt concentrations increased slowly with time. Increasing the concentration of salt in soils during germination resulted in decreasing germination followed by an increase in percentage of plant mortality (1, 22). Removing the salt at any time during the germination period, restored normal growth rates (29). Seeds of sugarbeets (Beta vulgaris), which failed to germinate but remained viable in a saline soil during an entire summer, germinated to produce normal seedlings the following fall when rains leached the salts from the vicinity of the seeds (18). Most plants seem to be more sensitive to salt during the seedling stage than during other stages of growth. 10 This may be because the tissues are tender and roots are shallow (40). Dumbroff and Cooper (25) found that osmotic stress was most deleterious to tomato plants (Lycopersicon esculentum) when applied during early growth, especially during the succulent seedling stage. Rice plants were found to be much more sensitive during the seedling stage than during germination at a given salinity level (44). Reports on other crops all support this fact. The variation in tolerance to salt during germination and seedling stages is insignificant in practice especially under high salinity levels where plants can make it through the germination stage, but fail to pass the seedling stage (44). Excess salinity reduces both rates of growth and total plant size. Forage and seed yield are usually reduced (40, 1). Increasing salinity levels in soybeans (Glycine max) increased plant mortality, and leaf necrosis. It also reduced green leaf color, leaflet size, dry stem pro- duction, plant height, seed yield and decreased seed quality, as would be expected as excess salinity increased accumulation of chloride in stems and leaves (1). Growth of most plants tested under saline conditions showed similar responses except for leaf color which darkened under increasing salinity, in the case of barley (9), alfalfa (l9) and beans (30). ll The degree of salt tolerance differs widely between plants. Barley was found to be a salt tolerant crop even at high salt concentrations. Ayers and Hayward (9) reported that "California mariout" barley germinated fairly well at high salt concentrations. In addition, they found that sugarbeets germinated poorly, and kidney beans did not germinate at all at moderate salt concentrations. Similar- ly, George and Williams (31) also found that "California mariout" germinated at higher salt concentrations compared to strawberry clover (Trifolium fragiferum) and Ladino clover. In other tests, barley was als‘ .snd to be more tolerant than wheat (42) and oats (Avena sativa) (10). Plants may have different degrees of salt tolerance during their growth stages (12). Sugarbeets are very tolerant to salt during the latter stages of growth, but are extremely sensitive to salt during germination. On the other hand, barley was found to be tolerant of salt during all stages of growth, although it was more sensitive during germination than at later stages (7). Barley plants grown under saline conditions have shown a decrease in vegetative vigor as evidenced by shortening of stems and decreased straw weights while maintaining essentially full yields of grains (9). Rice (Oryza sativa) plants were found to be more tolerant of salt during germination than 12 during the seedling stage (2, 3, 38, 44). Tolerance of rice seedlings to salt showed an appreciable increase at six-weeks of age (39). This increase in tolerance seems to cease later. The same study showed that these plants developed essentially normal straw yields but produced little or no grain. The degree of salt tolerance differs not only between plants, but also among species and varieties. Wheat varieties showed different responses at different levels of salinity during germination (11). A decrement of 50% in germination of a sensitive variety was found at a salinity level of -16 atm., while the tolerant variety showed the same decrement but at a salinity level of -20 atm. In tomatoes, Rush and Epstein (46) indicated that Galapagos ecotypes were more tolerant than the esculentum cultivars. Abel and Mackenzie (l), worked on soybeans, and noticed that tolerant varieties under saline conditions had little or no leaf necrosis, compared with other intermediate salt tolerant varieties. They then suggested the presence of inheritance factors for salt tolerance. Maddur (41) stated that salt tolerance in barley was carried out by partially dominant genes during germination and partial to complete dominant genes during early growth stages. Donovan (24) related differences in salt tolerance to the differences in 13 the imbibitional ability and a selective permeability of the seeds and/or the coleoptile epidermis to salt. Hunt (37) worked on intermediate wheatgrass and found that salt tolerance was a highly heritable characteristic. He found a great deal of variation between clones during germination and seedling stages. He also found that selection within species of tall wheatgrass produced greater salt tolerant strains. In another study, Dewey (22) also stated that selection in salt tolerance could be effective on crested wheatgrass (Agropyron cristatum) to obtain a salt tolerant strain. However, selected wheatgrass plants from germination tests did not show an appreciable improvement when planted in the field (23). F1 hybrids of two varieties of rice showed high resistance to salinization when compared to their parents for number of spikelets per panicle, panicle weight and grain yield per plant (4). The character of salt tolerance can be found in the wild relative species of some crops. A successful attempt was made to introduce this characteristic to the cultivated tomato plants, from the Galapagos salt—tolerant wild species L. cheesmanii (28). A salt tolerant plant species or variety does not mean that it will tolerate salt throughout it's life just because 14 it has a good tolerance during the germination period (8). Hunt (37) found little or no correlation between salt tolerance of intermediate wheatgrass during the seedling stage and later growth stages. Abel and Mackenzie (1) also noticed in soybeans that salt tolerance during germination and in the later stages was not apparently related. Barley cultivars showed different responses throughout their life cycle (43). One cultivar coped with high salt stress at emergence but did not make the transition into the vegeta- tive phase under high stress. Another cultivar, on the other hand, showed the opposite reaction. This may suggest the presence of two sets of genes for each stage. 15 MATERIALS AND METHODS Six varieties of barley previously selected by Maddur (41) according to their degree of tolerance to salt at germination stage. Their local names, identification, source and degree of tolerance to salt are given in Table 1. TABLE 1. Barley varieties that were used in this study. Varietnyame Identification Source Degree of Tolerance California Mariout CI 1455 Egypt Tolerant Lajbjey Drosihezy A not available Denmark Tolerant Coho CI 13852 USA Moderate Ingrid CI 10083 Sweden Moderate Mashu Mugi CI 11226 Japan Sensitive Orge Saida 183 not available Algeria Sensitive I. Screening test at germination stage: A screening method similar to the one developed by Whitmore and Sparrow for Laboratory malting (50) was applied with some modifications to fit the purpose of this study. Seeds were tested against 16,000 ppm.and 20,000 ppm NaCl. These levels were found to be critical for barley seeds at germination. The salt solutions were prepared by dissolving an equivalent amount of table salt (Iodine free) in a proper 16 volume of distilled water. Twenty-five seeds from each variety were placed in a 15 x 1.7 cm test tube. A volume of 15 cc of solution was poured into the test tubes. The test tubes were then placed in the growth chamber at 12°C for 48 hours. The solutions were changed every 12 hours. At the end of 48 hours, the solutions were filtered off and the kernels mopped to remove excess moisture. They were then placed back in their test tubes and stoppered with porous foam rubber. The test tubes were placed in the growth chamber at 17°C for 6 days. On alternate days the germinating seeds were carefully removed from the test tubes, to prevent rootlets from tangling together. This test was repeated three times. II. Mature-Plant Test: This test was made to determine the degree of tolerance of barley plants during later growth stages. The same varieties of barley mentioned in the germination test were used. The seeds were taken from plants grown the previous season. Plants were grown in pots, using sandy loam soil. California Mariout, a known salt tolerant variety, was used as a check variety. Two seeds of each variety along with 17 two seeds of California Mariout were planted in each pot. Three concentrations of sodium chloride (iodine free table salt) were used, 0, 16,000 and 24,000 parts per million. A total of 15 pots per replication were employed and the experiment was replicated three times. Pots as well as replications were randomly distributed and placed on three individual benches. The benches were about 30 cm high and wide enough to leave space among pots for better illumina- tion. The experiment was conducted in the growth chamber, where the temperature was kept at 18°C during day hours and 10°C during night hours. A total of 16 hours of light were received by the plants daily. Seeds were sown on April 30, 1978. Treatment solutions were applied on the 14th of May when the plants were in the three to four leaf stage. Each culture was to receive about one third of a liter of the solution on an alternate day. On the other days, the cultures were flooded with water to prevent salt accumulation. A complete fertilizer was applied to the plants once a week. Pots were rotated every week within and between replications. On the 28th of July, 1978, plants were harvested for measurements. The harvesting was done at the soil surface so that only the upper portions of the plants were involved 18 in the readings. Fresh weight, height, tiller number, and the dry weight of the plants were taken. Measurements on plant roots were ignored due to the difficulties in obtain- ing the roots of each individual plant. Yield was also ignored since the varieties had different maturity dates. III. Diallel Cross: To study the inheritance of salt tolerance in barley plants during the germination stage, a 6 x 6 diallel cross was made. Diallel cross analysis by Jinks (37) and Hayman (33) was employed in this study since salt tolerance was found to be a continuous rather than discrete variable (41). All possible crosses, including selfing between the selected parents were made with the assumption that: l) the parents were homozygous, 2) the inheritance was diploid, 3) genes at different loci were independently distributed in the parents, 4) no multiple allelism, 5) absence of maternal effects. In the analysis, the following second degree statistics were calculated: 1) the variance of parents (Vp) 2) the variance of the offspring of each parental 19 array (Vr) and, 3) the covariance of the offspring of each array with the non-recurring parent (Wr). The regression of Wr and Vr was obtained and Vr was plotted against Wr. Consistency of (Wr - Vr) over arrays and the significance of the regression of wr on Vr should jointly indicate the validity of the hypothesis postulated. Consistency of (Wr - Vr) was tested by using the formula: t = /£ig (Var Vr - Var Wr)2/Var Vr x Var Wr - cov2(Vr.Wr) with r—2 degrees of freedom, r being the number of parents. Significance of t indicated failure of the hypothesis. Significance test of the regression of Wr on Vr was carried out by the formula: _ b — 0 = l - b t1 — Sb and t2 _—Sb_ where Sb = /Szy.x/ZX2 with r-2 degrees of freedom. Non- significance of t1 indicated failure of the hypothesis, while significance of t1 indicated the presence of dominance. The significance of t2 indicated that non-allelic gene interaction was present. On Mendelian grounds, the array of offspring of the most dominant parent would be the least variable array and should have the smallest variance and covariance. The 20 opposite would be true for the array of offspring of the most recessive parent. The parabola wr2 = Vp Vr, delimited the area in which coordinate data (Wr, Vr) must occur. The line of unit slope (b=l) through the origin and Vr,Wr (where Wr was the mean of the covariances and Fr the mean of the variances) was the line of complete dominance. Move- ment of the regression line of unit slope upward relative to the line of complete dominance would denote partial dominance, while movement downwards would denote overdomi— nance. Non-allelic interaction, if present, would move the line to the right and drop its slope below the expected value of unity. The diallel cross was made among six parental varieties mentioned earlier in this chapter. The fifteen crosses of the 6 x 6 diallel were made in the greenhouse in the winter of 1977. F1 seeds were grown in the greenhouse to obtain F2 seeds. F2 seeds were then grown in the field to obtain F3 seeds. The study material was confined to the F3 seeds of the 15 crosses due to the low amount of F2 seeds. The F3 seeds of the 15 crosses along with the six parents were tested for salt tolerance at the germination stage in a 16,000 ppm NaCl solution as described earlier. 21 RESULTS 1. Mature plant test: These results indicate that the salt concentration gradient had a depressing effect on the dry weight of the six varieties studied. Increasing salt concentrations significantly reduced the growth of the plants as represent- ed by the dry weight (Figure l). The regression analysis in Figure 1 indicates a strong linear relationship between dry weight and salt concentra- tion. The analysis of variance (Table 2) for the dry weight and the regression analysis show significant differences between the six varieties. TABLE 2. Mean squares of salt tolerance scores as represent- ed by dry weights of the six varieties of barley at maturity. Source of Degrees of Mean Square F Variation Freedom Blocks 2 25.963 Entries 5 100.788 7.185** Error (a) 10 14.028 Treatment 2 1202.884 ll8.69** Entries x treatment 10 35.236 3.477** Error (b) 24 10.134 ** P i 0.01 22 The regression lines in Figure 2 show that there are two kinds of behavior: varieties Orge Saida 183, Coho, and California Mariout have a similar performance and may be considered salt tolerant, while other varieties showed sensitivity to salt. Differences between regression lines of the six varieties showed different levels of significance. A significant difference (p <.05) was found between Lajbjey Drosihezy A and Orge Saida 183, California Mariout, and Coho (Table 3). There were no significant differences among the last three varieties. Regression lines of the varieties Lajbjey Drosihezy A, Ingrid, and Mashu Mugi show different patterns, but all have greater negative response when compared to the rest. Lajbjey Drosihezy A seems to be the most affected by salt concentration. Ingrid and Mashu Mugi show similar reactions to salt. Fresh weight has also shown a strong linear relation- ship with salt concentration (Figure 3). Differences between regression means were not significant, although regression lines of the six varieties show somewhat differ- ent patterns (Figure 4). Varieties Ingrid and Mashu Mugi appear to be more depressed than the others. Lajbjey Drosihezy A lies close to Orge Saida 183, Coho, and 23 .mmfiuofium> Amaumo o «o uswwoz hum HmuOu ocu co cowumuucmocoo Homz mo uomwmm I H .on .0a2 2mm ooo¢~ coca. o .m .09 .mp .om mooooo.ol u n mm 'lM A80 (SWVUO) 24 mmoooo.on oomooo.0I oqqooo.OI qmqooo.o: oawooo.on N©HHOO.OI 3.0.0.0433 .mofluofium> mmfiumo e we unwfiw3 hue ozu :o coaumuucmocoo Homz mo uuowmm I N .052 Sam “ms: :cmmz usowumx «Houomwamo me meamm mwuo osoo efluwco < zuwzamoua zonnmmq HNMQ’MO 0v 'lM AUG (SWVHD) 25 TABLE 3. t values for the difference between two slopes for the six varieties of barley for dry weight, fresh weight, height and tiller number. Varieties Dry Weight Fresh Height Tiller Weight Number L. Drosihezy A and Ingrid 2.958+ 0.33 0.27 1.106 Coho 6.12** 0.245 0.79 3.247* O. Saida 183 6.068** 0.39 0.94 3.558* C. Mariout 5.564** 0.22 0.682 2.09 Mashu Mugi 4.278* 0.49 0.02 2.58 Ingrid and Coho 3.16* 0.57 0.79 2.14 0. Saida 183 3.12* 0.723 1.2 2.45 C. Mariout 2.6 0.111 0.952 0.98 Mashu Mugi 1.32 0.1629 0.29 1.476 Coho and 0. Saida 183 0.05 0.148 0.146 0.312 C. Mariout 0.5 0.46 0.109 1.156 Mashu Mugi 1.84 0.74 0.77 0.664 0. Saida 183 and C. Mariout 0.5 0.65 0.256 1.467 Mashu Mugi 1.79 0.923 0.92 0.97 C. Mariout and Mashu Mugi 1.29 0.274 .663 0.492 t P i 0.10 * P i 0.05 ** P 3 0.01 26 .mofiuowum> moapme 0 mo uawwoz ammuw HmuOu m:u co coHumpucoucoo Huwz mo uoommm a m .uHm .052 Eng ooofw 0006— o .o~ .OV .0m .0m «QMHOOAVI n a (SWVHS)'1M H8383 27 .mowuowum> zmaumn o no unwwmz :mouw msu so cowumuucooooo Homz mo uomwwm .OmZ 2mm ooo¢~ 0006. c .UHm q ‘ mmfimaoo.ou n n own: same: .0 omoofioo.o- u n uaofiumz masuomaamo .m ommwooo.o- u a me mefimm mwuo .q mmhoaoo.o: u a once .m mmmoaoo.ou u a wfiumco .N msamaoo.o- u n < snmewmooo smnnnmo .H 'lM H8383 (SWVHS) 28 California Mariout in terms of their regressions. The analysis of variance is summarized in Table 4. TABLE 4. Mean squares of salt tolerance scores as repre- sented by fresh weights of the six varieties of barley at maturity. Source of Degrees of Mean Square F Variation Freedom Blocks 2 105.951 Entries 5 1137.813 41.86** Error (a) 10 27.182 Treatment 2 6378.513 109.1943** Entries x treatment 10 112.472 l.925+ Error (b) 24 58.414 + P i 0.10 ** P i 0.01 Height of the six varieties also decreased as a function of salt concentration as shown in the strong linear relation- ship (Figure 5). No significant differences between regres- sions were found (Table 3). Regression lines of the varieties Orge Saida 183, Coho, and California Mariout have a similar behavior as shown in Figure 6. Heights of Ingrid, Mashu Mugi and Lajbjey Drosihezy A, were regressed in a similar manner, but showed more depression than the first group. 29 .mmwumfium> Awaken 0 mo uswwms Hmuou men so coHumoucmocou Homz mo uowmwm I m .UHm sz 2%. ooodm 0000. o .3 ,0m 0300.0: u o 100— 30 1H9|3H (W0) .wowumwum> mmauwn 0 00 unwwo; msu co coaumuucmocoo Homz mo uommwm I 0 .on .Oaz 2nd ooo€~ 0000. o ooommoo.OI ooooaoo.OI oowwooo.0I camaooo.0I oomqaoo.OI mnmmaoo.OI II. n own: semmz .0 III/I / a Sachem: machoonamo .m [III a me meamm owuo .q a oeou .m a ewewaH .N a H < zumcwwoum >mnnmma COP 1H9|3H (W0) 31 The analysis of variance (Table 5), however, showed no significant differences regarding the entry-treatment inter- actions. TABLE 5. Mean squares of salt tolerance scores as represent— ed by heights of the six varieties of barley at maturity. Source of Degrees of Mean Square F Variation Freedom Block 2 32.932 Entries 5 502.833 15.0677** Error (a) 10 33.372 Treatment 2 3807.315 67.1l** Entries x treatment 10 44.288 .781 Error (b) 24 56.736 ** P 5 0.01 A strong linear relationship between tiller number and salt concentration was also found (Figure 7). Figure 8 shows the effect of salt concentration on tiller number of the six varieties. Orge Saida 183 and Coho show superior performance under even the highest concentration (24,000 ppm). California Mariout and Mashu Mugi show some depression, while Lajbjey Drosihezy A and Ingrid were the most affected 32 .mowumwum> zmaumn 0 mo Hones: Hoaafiu Hmuou wsu co coaumuuaoocoo Homz mo uommmm I n .UHm .002 Sam oooem 0000. o Hoaooo.OI n 3 OF SHSWMl 33 .mowuowum> xmaumc 0 mo honesa umHHHu mnu :o seguMpucmoaoo H002 mo wooumm I w .UHm .0u2 Zan— oooem 0000— o .I III, -"I#"/ .I ’l’ "" ®" .I.’ [I’ll 'm.’ Ill- ”P'lol I.I“IIT‘1'II'IIIII'IIIVIII I. 4’. L11, I I. ,I ”I, _.| I.I III 10. 1 0’0, I’ll, m. .I. III, ’.’.’ IIan S .I. [IA ’0 I. I.I I.I. .m— I F]. I. OOOHOOO.OI u 3 Hwaz SSmNE .0 «mmH000.0I u o uaoaumz manu0mwamo .m .0N msooooo.o- u a mum «swam mweo .s ammoooo.OI u e 0:00 .m ommuooo.OI u n kuwfiH .N H00m000.0- u a < sumefimopa smegma; .H 34 by salt concentration. The analysis of variance is summarized in Table 6. TABLE 6. Mean squares of salt tolerance scores as represent- ed by tiller number of the six varieties of barley at maturity. Source of Degrees of Mean Square F Variation Freedom Block 2 2.905 Entries ' 5 58.716 24.98** Error (a) 10 2.350 Treatment 2 66.812 15.15** Entries x treatment 10 8.156 1.849 Error (b) 24 4.410 ** P i 0.01 The average moisture content (grams) per gram of dry matter of the above ground parts for the six varieties at the three concentrations was calculated and shown in Table 7. Generally, the six varieties had more moisture per gram of dry matter in the 16,000 and 24,000 ppm than in the control, except for California Mariout which did the reverse. Lajbjey Drosihezy A, seems to have more moisture per gram of dry matter at the 16,000 ppm and the 24,000 ppm than did 35 the rest. Ingrid and Mashu Mugi have shown a moderate accumulation of moisture in their tissues. Coho and Orge Saida 183 have shown the least moisture per gram of dry mat- ter. California Mariout absorbed less water per gram of dry matter in both the 16,000 and the 24,000 ppm than did the control. TABLE 7. Average moisture content (grams) per gram of dry matter of the six varieties of barley at three levels of salinity. NaCl Concentration Variety 0 ppm 16,000 ppm 24,000 ppm 1 Lajbjey Drosihezy A 1.12 3.10 4.19 2 Ingrid 1.85 3.45 3.24 3 Coho 2.88 4.08 4.64 4. Orge Saida 183 2.14 3.00 3.05 5 California Mariout 2.16 1.85 1.89 6 Mashu Mugi 2.33 3.09 4.10 The varieties Lajbjey Drosihezy A and Mashu Mugi show a proportional relationship between moisture content and salt concentration, i.e. moisture content increased as a function of increasing salt concentration. This is not true, however, for the rest of the varieties since they retained moisture at a similar level in the 16,000 and the 24,000 ppm. 36 2. Germination test: The germination test showed that increasing salt con- centration significantly decreased germination percentage of the six varieties of barley tested. The regression analysis (Figure 9) shows that there is a strong linear relationship between salt concentration and germination percentage. The six varieties of barley behaved differently to salt treatment as shown by their different regression lines (Figure 10) and the analysis of variance (Table 8). California Mariout was superior when TABLE 8. Mean squares of salt tolerance scores as repre- sented by germinating seeds of the six varieties of barley. Source of Degrees of Mean Square F Variation Freedom Block 2 1.241 Entries 5 119.529 23.91** Error (a) 10 2.085 Treatment 2 1203.574 309.5** Entries x treatment 10 25.619 6.5879** Error (b) 24 3.889 ** P i 0.01 37 .mmwumfium> amauwo 0 mo mwmucmouoa :ofiuQGHEuww Hmuou mnu :o coaumuuaooaoo Homz mo uomumm I 0 .on .002 Ema 0000a 0000. 0 C ‘ mmqomoo.0I u o 00— % NOIlVNIWHBE) 38 .mwfiumfipm> kmaumn 0 mo ammucmoumm :owu00HEuww wzu co coaumpucmucoo Homz we uoouum _002 Sam ooodN ooodp 0H .uHm 00m0000.0- u 0 «0:2 sewn: 0 onmmooo.o- u 0 uaofiumz maceoLHHmu m owfioaoo.0- u a 000 meamm mwco .s 0000000.o- u 0 oeou .m mmwm000.o: u 0 enemaH .N 0000000.0- u 0 < sumewmoua samamme .H % NOUVNIWUBO 39 compared to other varieties. Varieties Lajbjey Drosihezy A, Orge Saida 183, and Mashu Mugi were most affected by salt, while varieties Ingrid and Coho have shown an intermediate performance. According to their performance in the germina- tion test, California Mariout was considered salt tolerant; Ingrid and Coho were intermediate; Lajbjey Drosihezy A, Orge Saida 183, and Mashu Mugi salt sensitive. Differences among regression lines were calculated and showed different degrees of significance (Table 9). Highly significant differences were found between California Mariout and the salt sensitive varieties; while significant differences were found between the later lines and the intermediate ones. No significant differences, however, were shown between California Mariout and the intermediate lines. Correlations between tolerance to salt during the germination stage and the mature-plant stage were calculated (Table 10). The regression coefficients obtained for the six lines in the germination stage were correlated with those of dry weight, fresh weight, height, and tiller number in the mature-plant stage. The regression coefficients were used in this test because they represent an estimate for tolerance to salt. 40 TABLE 9. t values for the difference between two slopes for the six varieties of barley for germination. Varieties Lajbjey Drosihezy A and Ingrid .59 Coho .22 Orge Saida 183 .612 California Mariout .63+ Mashu Mugi .7 Ingrid and Coho .37 Orge Saida 183 .2+ California Mariout .04 Mashu Mugi .3t Coho and Orge Saida 183 .83 California Mariout .41 Mashu Mugi .93 Orge Saida 183 and California Mariout .24* Mashu Mugi .093 California Mariout and Mashu Mugi .34* + P i 0.10 * P 3 0.05 41 TABLE 10. The correlations among regression coefficients for germination % and mature-plant characteristics. Stage of Growth d.f. r Germination and dry weight 4 + .183 Germination and fresh weight 4 - .061 Germination and height 4 + .371 Germination and tiller number 4 - .158 The calculated r values for germination and dry weight and germination and height were positive while they were negative for fresh weight and tillers. These values, however, were not found significant, indicating no signifi- cant correlation between salt tolerance in the two stages of growth. 3. Genetic investigation: The genetic investigation will be discussed on the basis of the results of the salt tolerance tests in the germination stage on the F3 progenies of the 6 x 6 diallel set. The diallel cross data is required to show a signifi- cant variation among hybrids for the test to contain reliable genetic information. The step is usually examined prior to carrying on further analysis. 42 The results of the analysis of variance of the fifteen F3 hybrid progenies for salt tolerance test at the germina— tion stage is presented in Table 11. A highly significant TABLE 11. Mean squares for salt tolerance scores in the germination stage of the F3 of the 6 x 6 diallel cross set. Source of Degrees of Mean Square F Variation Freedom Block 2 11.822 Entries 14 47.876 6.353** Error (a) 28 7.537 Treatment 2 3817.356 655.9** Entries x treatment 28 13.998 2.41** Error (b) 60 5.820 ** P i 0.01 difference existed among entries. Consequently, the genetic relationship among this set of selected parents and progenies was analyzed using the technique of the Jinks-Hayman diallel cross analysis and the graphical analyses were based on the variance and the covariance of the arrays. The F3 data of salt tolerance in the germination stage is summarized in Table 12. Each value is the average germination percentage of 75 F3 seeds. The array's variance 43 (Vr) and covariance (Wr) are shown on the right hand side of the table. The table shows that the means of the crosses do not consistently lie in the range of the parents. Crosses l x 2, 2 x 4, 2 x 6, 3 x 4, 3 x 6, 4 x 5, and 5 x 6 have a mean value in the range of their parents. Cross 1 x 6 was the only combination that showed transgressive effect since the mean was higher than the mean of either of its parents. The mean values of crosses 1 x 3, 1 x 5, 2 x 3, 4 x 6 were close to that of parents 3, 1, 2, and 4, respectively. Crosses that showed mean values less than their lower parents were 1 x 4, 2 x 5, and 3 x 5. The Vr values show that array 3 was the least variable and array 5 the most variable. The t values for the test of the consistency of the variable (Wr - Vr) over arrays was calculated at t = 1.66 and found to be not significant (P > .90) indicating the validity of the postulated hypothesis. The regression graph of Wr on Vr is shown in Figure 12 along with the limiting parabola Wr2 = Wer. The graphical analysis shows that Wr and Vr enjoy an almost linear relationship with the regression coefficient b = 1.533** significantly greater than zero but not more than one. 44 n00.0mm 0~H.m~N n0.sm ~0.~0 00.0w 00.00 00.00 00.00 00:: seam: 0 n00.00N 0H0.wmm 50.05 50.0m 00.n0 mm.0q 00.00 uaowumz .0 m 000.0HH N00.~0N mm.a~ 00.0w 00.~m mm.na 00H 00000 .0 q Ham.mm H00.00H mm.a0 00.Nm ~0.N0 0:00 m 000.000 an.NNN 00.00 00.00 eauwaH N 000.m0~ NN~.00~ 00.00 < sumeamoua .0 H HonEbz 93 H> 0 m N H mucmpmm Hmucoumm .umm mmouo Hmaamwm 0 x 0 map mo mm onu mo mmmeCmouoa cowumcwauow owmuw>< .NH mqm