Wide and Intra-specific Variation in Total Seed Protein in Chickpea ( Cicer arietinum L .) Hybrids Through SDS-PAGE

In the present study, 30 accessions of chickpea were analyzed for total seed protein profile using Sodium Dodecyl Sulphate-Poly Acraylamide Gel Electrophoresis (SDS-PAGE) to ascertain the extent of genetic variation. Based on dendrogram of SDS-PAGE, genotypes were divided into nine clusters taking 75% similarity as standard. From the clustering pattern of the 30 genotypes, it was clear that cluster III evolved as large cluster comprising 12 genotypes and clusters I, II, VI, VII, VIII and IX comprised single genotype each. Cluster IV and V comprised two and ten genotypes, respectively. The minimum similarity coefficient was 0.11 hence the genotypes, PG 061 and PG 059 were most dissimilar with PG041. It was observed that four genotypes namely PG 047, PG 052, PG 058 and PG 057 had shown similarity coefficient 1.0. Therefore, these genotypes may be considered genetically similar. SDS-PAGE can be used for characterization of cultivars and species, establishment of phylogenetic relationship among taxa and for genetic studies in many crops.

Pulses are the main source of vegetable protein in human consumption and occupies a wide range of agro climatic conditions and it contributes about 25 per cent to the global production (Anonymous, 2005). Chickpea play an important role in rainfed agriculture by improving physical, chemical and biological properties of soil and is considered an excellent crop for natural resource management, environmental security, crop diversification and consequently for viable agriculture. In the crop breeding programme to develop a suitable variety, breeder has to combine different genotypes with desirable characters into one variety. In order to identify true-recombinations, generally morphological markers are used. However, sometimes morphological markers may be influenced by the environment and are usually dominant or recessive and sometimes it is also difficult to find appropriate morphological marker.
In the recent years, SDS-PAGE markers have been used extensively for characterization of cultivars and species, establishment of phylogenetic relationship among taxa and for genetic studies in many crops. SDS PAGE is a valid technique and is increasingly being used an approach for species identification. Each variety or a group of varieties has specific protein banding patterns and on the basis of these patterns, they can be identified accordingly. Thirty accessions of Cicer arietinum (Chickpea) germplasm were analyzed for total seed protein profile using Sodium Dodecyl Sulphate -Polyacraylamide Gel Electrophoresis (SDS-PAGE) to ascertain the extent of genetic variation for total seed protein among the genotypes derived from wide and intervarietal crosses of chickpea based on SDS-PAGE.
Experimental material of the present study consisting 30 genotypes including 25 genotypes selected from segrregating generations of wide and intervarietal crosses (Table 1) and five released varieties of chickpea. Thirty entries were planted in a randomized complete block design with three replications during the crop season 2005-07. Row length was kept at 4 m and the spacing between two rows at 30 cm. Recommended package of practices was followed for raising the crop. Genetic relationship of progenies of a wide cross was also studied on biochemical basis using SDS-PAGE as per the standardized procedure for seed protein in chickpea (Singh et al., 1991). The stacking and separating gels, staining and destaining solutions were prepared according the protocol. Jaccard's similarity coefficient was used for calculating for the genetic similarity between progenies and parents (Jaccard, 1908). If this value is 1.00 then progenies are genetically same. If the value between two progenies is 0.00, then the progenies are genetically different.
In the present investigation, proteins were extracted from seeds of 30 chickpea genotypes and then separated on the basis of size and electrical charge in a gel matrix by SDS-PAGE. Distribution of 30 genotypes into different clusters is presented in Table 2. The banding pattern of genotypes is presented in Fig.2 and Fig. 3 and Table 3. All 30 genotypes were divided into nine clusters taking 75% similarity as standard based on electrophoretic banding pattern (with respect to number of bands, their thickness and intensity) of SDS-PAGE. Table 2 showed the cluster number and names of genotypes included in each cluster and dendrogram is represented in Fig.1. Cluster III was evolved as a large cluster comprising 12 genotypes. Clusters IV and V comprised of two and ten genotypes, respectively and clusters I, II, VI, VII, VIII and IX comprised single genotype each.   (Table 2). This may be due to differential recombination of the genes from parents into the different segregating generations. Also this may be due to resolution of limited number of loci for differentiation the genotypes for seed protein electrophoresis (Nisar et al., 2007). The genotypes, PG 055 and PG 056 separated into two different clusters with 75% similarity, inspite of having common parentage (BG362 x Avrodhi).
The minimum similarity coefficient was 0.11 hence the genotypes; PG 061 and PG 059 were most dissimilar with PG041 (Fig. 1). The parents of PG 041 were PG92-4 × Avrodhi and the parents of PG 061 and PG 059 were PG92-97 × C. reticulatum. Based on similarity coefficient of genotypes, it was observed that four genotypes namely PG 047, PG 052, PG 058 and PG 057 had shown similarity 157 coefficient 1. Therefore, these genotypes may be considered genetically similar (on the basis of SDS-PAGE). For this reason, all the other 24 genotypes had shown same similarity coefficient with these four genotypes (PG 047, PG 052, PG 057 and PG 058) separately (Hameed et al., 2009). SDS-PAGE analysis can be used to find out the wild progenitor of different crop species, existence of genetic diversity in germplasm and identification of germplasm and varieties (Ghafoor et al, 2002;Ghafoor et al., 2005 andSingh et al., 2004). Protein electrophoresis is also a powerful tool for population genetics (Parker et al., 1998) and the SDS-PAGE technology is particularly considered as a reliable way because storage proteins are largely independent of environmental fluctuations (Javid et al., 2004 andIqbal et al., 2005). Therefore, it is concluded that SDS-PAGE analysis is useful tool for a plant breeder for selection and identification of genotypes within a short period of time with greater efficiency.