PCR-RFLP analysis of growth hormone gene in extreme groups of colored broiler parent lines

V.K. Saxena, A.B. Pramod, K.A. Ahmed, N. Ravi, M. Saxena,A.K. Dev Roy and R.V. Singh
Central Avian Research Institute, Izatnagar - 243 122

Candidate gene analysis is one of the advance molecular approaches for understanding the genetic mechanism of a physiological phenomenon and also to find out a suitable molecular marker to be utilized in MAS.Chicken Growth hormone (cGH) gene located on chromosome 1 an important gene involved in somatic growth. It is 3.5 kb in size and consisting of 5 exons & 4 introns. cGH gene is associated with various production traits e.g. body weight, egg production, carcass traits, fatness and leanness, immune-responsiveness etc. Intron IV of cGH is highly polymorphic therefore chosen as candidate gene for analysis in broiler chicken parent lines Using specific reverse and forward primers the intron IV region of cGH was amplified as 1164 bp band in individuals of two groups of coloured synthetic male line (CSML) i.e. one group having extremely higher body weight and the other having extremely low body weight at 5 weeks. The reaction mixture (25μl) contained: Buffer-1 X, Taq polymerase 2 U, dNTPS-0.2mM, Primers- 20 pmoles each, DNA template-100ng. The optimized cycling conditions were: Initial denaturation: 94ºC for 4 m; 35 cycles with denaturation at 94ºC for 30s, annealing at 62ºC for 2 m, extension 72ºC for 1.5m and final extension was at 72ºC for 5m. The amplified products from individual samples were column purified and subjected to digestion with HaeIII and TaqI restriction enzymes as per recommendations of the manufacturer of the enzymes. The digested products were visualized on PAGE stained by silver staining. PCR-RFLP with HaeIII showed monomorphic pattern in both groups. All the individuals analyzed revealed bands of 480, 240, 215, 130 and 80 bp. However, TaqI revealed polymorphism between the two groups. A total of three genotypes i.e. AA (1164/1164 bp), AB (1164/513, 651 bp) and BC (513, 651/ 440, 651, 73 bp) were observed. The genotypic frequencies of the AA, AB and BC genotypes in high body weight group in high body weight group were 0.25, 0.5 and 0.25 respectively. Corresponding values in low body weight group were 0.50, 0.0 and 0.50. The preliminary results revealed encouraging findings for analyzing the cGH and also other candidate genes related to growth more extensively.

Source : IPSACON-2005

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