الفهرس | Only 14 pages are availabe for public view |
Abstract Faba bean (Vicia faba L.) is an annual, diploid (2n=2x=12), widely distributed species and is considered to be the first leguminous crop that is grown in Egypt. Total yield is consumed as a vegetable either green or dried seeds in human feed hence it has high levels of protein (30% on average), carbohydrates (58%), fiber, vitamins and minerals. In Egypt, the total production of faba bean is still insufficient to cover the local consumption (about 28% of the local consumption). So, there is a great need to overcome this gap between total production and local consumption by: 1- expanding the cultivated area of faba bean through the new reclaimed lands; where water, salinity and heat stress and so forth are the important difficulty, as well as, 2- increasing the vertical production through introduction of new varieties with high yield potential and the ability to withstand different environmental stresses. This study was carried out to evaluate the performance of seven divergent faba bean genotypes that included: four local cultivars (Misr-1, Misr-3, Nubaria- 2 and Sakha-4), two newly released lines (NBL-Mar.3 and NBL-5) and one exotic genotype from ICARDA (L3) under two conditions; prevailing terminal heat stress conditions of the New Valley and adequate conditions of Ashmoun, El-Menofia governorate during the winter growing season (2012/2013). Several parameters were used to compare between the studied divergent seven faba bean genotypes: (1) Yield traits. (2) Stress tolerance indices based on yield performance of the tested seven faba bean genotypes under stress and non-stress environments. (3) Biochemical markers as revealed by: a- Polyacrylamide gel electrophoresis of seed and leaf protein b- Four isozyme systems. (4) Molecular markers and fingerprints analysis using two techniques: a- Inter simple sequence repeats polymerase chain reaction (ISSR-PCR). b- Inter-retrotransposons amplified polymorphism (IRAP-PCR).(5) Using ISSR and IRAP generating alleles for testing the existence of genotypes variability by calculating polymorphic information content (PIC) and discriminative power (D). (6) UPGMA cluster analysis of the seven genotypes tested based on the ISSRPCR and IRAP-PCR data by estimating genetic variability and analyzing the relationships among the seven genotypes. The yield traits of plant height, number of primary branches per plant, number of pods per plant, number of seeds per pod, number of seeds per plant, seed index (100-seeds weight g) and seed yield per plant(g) were measured for the seven faba bean genotypes tested under the two experimental locations. Significant differences were recorded among the seven genotypes for all measured characters at both locations which may due to their different genetic background. The stress conditions at New Valley have resulted in a reduction of yield components and its attributes for all genotypes; this was illustrated by the mean readings of the studied traits comparing between the two locations. Superiority of Misr-1 genotype at Ashmoun (non-stressed) was due to its relative advantages in number of primary branches/plant, number of pods/plant, number of seeds/plant and seed yield/plant. Contrary, superiority of Sakha-4 genotype at the New Valley (stressed) used to be as a result of its relative advantages in plant height, number of primary branches/plant, seed index (weight of 100 seeds in g.) and seed yield/plant. These results reflected the importance of these traits as effective yield contributors and should be used as criteria for selection in future faba bean improvement program. Regarding seed yield/plant (g), it could be concluded that the genotype Sakha-4 (which ranked third place under adequate conditions while ranked first under stress conditions) is the most tolerant genotype that it was adaptable for the prevailing condition of New Valley and gave the highest level of yielding capacity. Otherwise, Nubaria-2 is the least tolerant one, which ranked last genotype under both non-stress conditions as well as stress conditions. The previous superior genotype i.e. Sakha-4 can be exploits in faba bean improving programs. Seed yield/plant (g) differences among the tested genotypes under both locations reflect a fluctuation response in each of the used eleven tolerance indices. This confirmed by detecting the lowest value of SSI index as well as the highest values of GMP, HM, STI, k2STI, YI and YSI indices for Sakha-4 genotype. Therefore, Sakha-4 genotype could be considered as the most heat stress tolerant genotype. On the contrary, Nubaria-2 genotype (which ranked last genotype under non-stress conditions as well as under stress conditions) achieved the highest value of SSI index and the lowest values of MP, GMP, HM, STI, K1STI, K2STI, YI, and YSI indices, therefor, Nubaria-2 genotype could be considered as the least tolerant one. Thus, this investigation suggested that the results of SSI, GMP, HM, STI, K2STI, YI and YSI indices in selection of genotypes are appropriates for selection of heat tolerant/susceptible genotypes, and therefor to determine the most appropriate heat tolerance indices of faba bean genotypes. Ployacrylamide gel electrophoresis analysis of seed protein patterns revealed forty six bands under adequate location of Ashmoun. Such bands were detected at approximately molecular mass ranging between 107.82 and 8.61 kDa, with polymorphism of 93.48%. Eleven unique bands represented as (M+) were recorded. These unique bands could be used as positive biochemical markers (M+) for the faba bean genotypes under study. Also, 12 polymorphic bands are appeared in all studied genotypes except one, these bands are represented as (M-) and could be used as negative biochemical markers for the faba bean genotypes under study. While, forty four bands were recorded in seed protein patterns under stressed location of New Valley. Such bands were detected at approximately molecular mass ranging between 148.57 and 4.88 kDa, with 59 % polymorphism. Four unique bands represented as (M+) were scored. These bands could be used as positive biochemical markers (M+) for heat stress for the faba bean genotypes under study. Also, 10 polymorphic bands are appeared in all studied genotypes except one; these bands are represented as (M-) and could be used as negative biochemical markers for the tested faba bean genotypes under heat stress. Ployacrylamide gel electrophoresis analysis of leaf protein patterns revealed thirty three bands under adequate location of Ashmoun. Such bands were detected at roughly molecular mass ranging between 359.67 and 16.45 kDa, with polymorphism of 81.8%. Five unique bands were found and represented as (M+). These unique bands could be used as positive biochemical markers (M+) for the faba bean genotypes under study. Also, 8 polymorphic bands were appeared in all studied genotypes except one, these bands were represented as (M-) and could be used as negative biochemical markers for the faba bean genotypes under study. Whilst, forty three bands were observed in the leaf protein pattern under stressed location of New Valley. Such bands were detected at approximately molecular mass ranging between 100.21 and 15.79 kDa, with polymorphism percentage of 65.12%. Seven unique bands were scored and represented as (M+). These unique bands could be used as positive biochemical markers for heat stress for the faba bean genotypes under study. Also, four polymorphic bands were appeared in all studied genotypes except one; these bands were represented as (M-) and could be used as negative biochemical markers for the tested faba bean genotypes under heat stress. Native polyacrylamide gel electrophoresis was used to detect isozyme fingerprints of the seven faba bean genotypes tested under both adequate and stressed locations and to detect biochemical markers for faba bean tolerance to heat stress. This was done using four isozymes including α- esterase, peroxidase, alkohol dehydrogenase and polyphenyl oxidase. In this concern, under adequate location of Ashmoun, a total of twenty one bands were recorded by the four isozymes patterns with 47.6% polymorphism. Different values of polymorphism were detected in the seven genotypes under study, ranging from zero % (α- esterase) to 100 % (polyphenyl oxidase). Peroxidase patterns gave two unique bands (Px3 and Px4) and scored in the NBL-5 genotype which could be used as biochemical marker for this genotype under non-stressed conditions. Also, among the polymorphic bands of all patterns two bands out of them i.e. Px2 and PPO1 were present in all genotypes except NBL (Mar.3) and L3 respectively, and could be used as negative biochemical markers for these genotypes. Otherwise, under stressed location of New Valley, a total of twenty five bands were recorded by the four isozymes patterns with 72 % polymorphism. Different values of polymorphism were detected in the seven genotypes under study, ranging from 33 % (α-esterase) to 100 % (peroxidase). Peroxidase patterns gave one unique band (Px3) and scored in the more tolerant genotype Misr-1 and could be used as a biochemical marker for this genotype under heat stress conditions. Also, polyphenyl oxidase patterns gave three unique bands (PPO4, PPO6 and PPO8), which scored in the least tolerant genotype Nubaria-2, therefore these bands could be considered as biochemical markers for heat stress susceptibility. Furthermore, two bands (Est3 and Est4) were shared polymorphic ones and existed in the two more tolerant and most tolerant genotypes Misr-1 and Sakha-4, therefore these bands could be considered as biochemical markers for heat stress tolerance. Ten oligonucleotide primers were used to establish ISSR-PCR fingerprints of seven faba bean genotypes. A total number of 85 amplicons (amplified fragments) were generated, with 88.2% polymorphism. The highest percentage of polymorphism (100 %) was detected with each of the primers B44, 17898B, HB- 10, HB-13, HB-14 while the lowest one (57.1 %) was recorded in the primer HB- 09. The ISSR-PCR fragments generated by using the ten primers could be used as genotypic specific markers. A total of 26 specific markers including either the presence (M+) or absence (M-) were identified out of the total amplified fragments in the tested genotypes, and they could be considered as marker assisted selection (MAS) for heat stress tolerance and as molecular fingerprints for tested genotypes. Values of polymorphic information content (PIC) and discriminating power (D) indicated that primer HB-14 was the most efficient to analyze genetic variability among faba bean genotypes with values of 0.99 and 0.889, respectively, followed by primers 17898B and HB-10. A whole of 141 amplicons (amplified fragments) were generated via the ten IRAP primers, with 87.3% polymorphism. The highest percentage of polymorphism (100 %) was detected with each of the primers (4314) and (4373), while the lowest one (50 %) was recorded in the primer (4362). The IRAP-PCR fragments generated by using the ten primers could be used as genotypic specific markers. A total of 60 specific markers including either the presence (M+) or absence (M-) were identified among the ten primers; such bands could be considered as molecular markers (marker assisted selection) for heat stress tolerance as well as molecular fingerprints for the tested genotypes. Values of PIC and D indicated that primer 4376 was the most efficient to analyze genetic variability with values of 0.988 and 0.935, respectively, followed by primers 4314 and 4374. UPGMA cluster analysis of the seven genotypes tested based on the ISSRPCR and IRAP-PCR data revealed that, Nubaria-2 (least tolerant) genotype represented as a separate phenetic line which had the lowest similarity values with all other genotypes. The other genotypes were classified into two main clusters. The first cluster included three genotypes (L3, NBL-5 and NBL-Mar.3); it is worthy to note that, these three genotypes include one common ancestor; hence, this confirmed the high similarity value between them. The second cluster included three genotypes represented by a separate phenetic line that comprises Sakha-4 (most tolerant) genotype and a sub-cluster comprises Misr-3 and Misr-1 genotypes, also it is worthy to note that Misr-1 genotype is one of the ancestors of Misr-3 genotype. |