GENETIC AND MOLECULAR ANALYSES OF NODULATION IN CHICKPEA (Cicer arietinum L.)

Abstract

A hallmark trait of chickpea ( cicer arietinum L.) is its ability to form root nodules and to fix atmospheric nitrogen in symbiosis with compatible rhizobia. Chickpea plays a vital role in natural ecosystems, agriculture, and agro- forestry, where its ability to fix nitrogen in symbiosis makes it an excellent settler of low-N 2 environments, and economic and environmentally friendly crop. Forty seven chickpea genotypes were procured from Nuclear Institute for Food and Agriculture (NIFA), Peshawar, Gram Research Station (GRS), Karak, Pakistan and International Crops Research Institute for the Semi-Arid Tropics (ICRISAT), India. Entire experiments of the reported project were carried out from 2006 to 2009 at Agricultural University, and NIFA, Peshawar except molecular characterization, which was accomplished at Ehime University Matsuyama, Japan. All genotypes were characterized for marker traits, quantitative parameters, nodulation and molecular markers (SSR). Highly nodulated and non-nodulated parents were picked and hybridized to study mode of inheritance of nodulation and its linkage with marker trait loci. The germplasm was also grouped as desi (pink flower, green with purplish tings stem and colored seed coat) and kabuli (white flower, green stem and white seed coat) types. Highly significant differences and high heritability estimates were recorded for days to 50% flowering, days to maturity, leaf area, number of leaflets leaf -1 , plant height, 100 seed weight, biological yield plant -1 and grain yield plant -1 in all the genotypes. Genotypes from NIFA and GRS were nodulated while genotypes from ICRISAT were Nod - . All genotypes also differed highly and significantly for number of nodules plant -1 . The genotypes NDC 5-S10 and NDC 4-20-4 xproduced the maximum nodules plant -1 . Highly significant response of rhizobium inoculation was recorded for nodules plant -1 and seed yield plant -1 . Interaction of genotypes with treatments classified NDC 4-20-1(16.66) as highly Nod+ and Karak 3 (33 g) as high seed yielder plant -1 . The maximum genotypic mean for nodulation and seed yield plant -1 was recorded for accession NDC 5-S10 (14.83) and Karak 3 (30.20) respectively. Inoculated genotypes exceeded control in treatment means both for nodules plant -1 (10.33 & 7.22) and seed yield plant -1 (14.40 g & 10.59 g). Molecular characterization of 47 genotypes was performed using 10 simple sequence repeat (SSR) markers. Eight of the 10 SSR markers were polymorphic. Number of alleles ranged from 2 to 16, with an average of 7.4 locus -1 . Polymorphic information content (PIC) values ranged from 0.227 to 0.876, with an average of 0.636. The average PIC was 0.582 in desi and 0.577 in kabuli genotypes, shows that both groups are distinct. Significant genetic differentiation was found between desi and kabuli genotypes by using Analysis of molecular variance (AMOVA) under stepwise mutation assumption (R ST = 0.239, P ≤ 0.001). Unweighted pair-group method using arithmetic averages (UPGMA) and Minimum-evolution method (ME) trees as well as Principal Coordinate analysis (PCoA) classified the accessions into 6 groups and all the 6 accessions could be clearly separated. Grouping was mostly the same in both the phylogenetic trees and PCoA, but the branching order differed greatly. Inheritance of nodulation was studied in two cross combinations i.e., ICC 19181(non-nodulated and dark green leaves) x NDC 5-S10 (nodulated and light green leaves) - Hybrid A and ICC 19181 x NDC 4-20-4 (Nod+ and light green leaves) - Hybrid B. Hybrid A, showed monogenic dominant inheritance, while hybrid B showed duplicate gene action for nodulation confirming that both Nod + genotypes are from different clusters. Both hybrids revealed monogenic dominant inheritance of light green leaf color. Linkage study revealed that loci for nod and leaf color resides on the same chromosome at the distance of 15 centi Morgan (cM) in genotype NDC 5-S10 while in genotype NDC 4-20-4 the two loci for nodulation exists at the distance of 26 cM and 15 cM from the locus of leaf color. The current research findings show significant diversity both at morphological and molecular levels, and valuable results regarding rhizobial inoculation, inheritance and linkage study of nodulation, which could play a vital role in future chickpea breeding programs.