Record Details

Molecular Characterization of Casein Genes and the Effects of their Polymorphism on Selected Production Traits in Buffaloes

KrishiKosh

View Archive Info
 
 
Field Value
 
Title Molecular Characterization of Casein Genes and the Effects of their Polymorphism on Selected Production Traits in Buffaloes
 
Creator Darshan Raj, G.
 
Contributor Govindaiah M.G.
 
Subject ---
 
Description The present investigation was aimed at studying the genetic variation in casein gene locus between the three buffalo breeds viz, South Kanara, Surti and Murrah and to assess the genotypic frequency of different haplotypes at different regions of the gene. Attempts were also made to relate the haplotypes with milk composition. High salt method was employed to isolate Genomic DNA from South Kanara (50), Surti (50) and Murrah (50) buffaloes. The quality and quantity of DNA were determined by spectrophotometer and agarose gel electrophoresis.
A 1267 bp fragment of 5'-noncoding regulatory sequence of uS2-casein gene, a 329 bp fragment of exon 7 of p-casein gene and a 350 bp fragment of exon 4 of x-casein genes were amplified by PCR using oligonucleotide primers standardized for Bos taurus species. The sizes of the amplification products were similar for all the buffalo breeds studied, indicating the conservation of all the three casein genes in buffalo breeds.
The PCR amplicons of uS2-casein gene of different buffalo breeds were subjected to restriction digestion by the enzymes MaelI and EcoRV to study the polymorphic sites. An intact 1267 bp fragment was visible after digestion in all the animals. Such types of restriction patterns have not been reported earlier in cattle. The -1084 and -317 bp polymorphic positions of uS2-casein gene studied by the enzyme MaeII were genotyped as 'TT' and 'AA', respectively. In contrast to the RFLP studies of uS2-casein gene, where enzyme did not produce any restriction fragments, differences were revealed upon sequence data analysis. The adjacent bases of -186 bp polymorphic site (-184 and -185 bp positions) were mutated, suggesting that the EcoRV has limited applications in identifying haplotypes at -186 bp position in buffaloes.
In all the breeds studied, a restriction pattern having fragments of 266 bp and 84 bp was noticed following digestion of the 350 bp fragment x-casein gene by the enzyme HinfI, suggesting BB genotype in all the animals. No variation in the mobility of the fragments was found when the PCR amplicons of US2, p and K casein genes of different buffalo breeds were subjected to SSCP analysis. The results of SSCP and RFLP analysis were identical with respect to monomorphic pattern for the casein genes studied. The pattern of nucleotide sequence in all the three breeds indicated monomorphic nature for the US2, p and K-casein genes.
The nucleotide sequencing of the amplified Bubalus bubalis US2 promoter region indicated that the product size was 1262 bp which is smaller to that of the cattle for the same set of primers and no polymorphism was found within and between the breeds studied for 10 variable sites. Sequencing of amplified p-casein gene showed cytosine (C) at the + 8267 nucleotide position, which was seen in genotype "AA" only. K-casein gene sequence data indicated cytosine (C) at the + 5345 nucleotide position, which was found in genotype "BB" only. The BLAST alignment of 1262 nucleotides of buffalo US2- casein gene showed 96 per cent and 97 per cent homogeneity with the Bos taurus and Bos grunniens uS2-casein gene casein, respectively. The alignment of 329 bp p-casein gene showed 98 per cent identity with the Bubalis bubalus gene sequence. The BLAST search for 350 bp K-casein showed 98 per cent identity with Syncerus caffer nanus species. Translation data of p-casein gene showed 109 amino acids with serine amino acid at 69th position in all the animals studied, confirming the homozygous nature of the genotype in all the animals. Translation data of 350 bp of K-casein gene yielded 66 amino acids with alanine being the 45th amino acid.
The milk components and milk yield parameters were not associated with buffalo casein genotypes due to their monomorphic haplotype. The monomorphic pattern of the three casein genes (Us2, p, K) revealed by PCR-RFLP, SSCP technique and sequencing data analysis may be a buffalo specific character. There was a significant difference between the nucleotide sequence of milk protein genes of Bubalus huhalis and Bos taurus animals which need to be further studied to assess their effect on milk constituents of the two species.
 
Date 2016-06-21T12:52:13Z
2016-06-21T12:52:13Z
2006-10-05
 
Type Thesis
 
Identifier http://krishikosh.egranth.ac.in/handle/1/67581
 
Language en
 
Format application/pdf
 
Publisher Karnataka Veterinary, Animal and Fisheries Sciences University, Bidar