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20113
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Subject: Animal Genetics and Breeding × End date: 2011 × Start date: 2010 ×
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    ThesisItemOpen Access
    ESTROGEN RECEPTOR GENE POLYMORPHISM IN VECHUR AND CROSSBRED CATTLE OF KERALA
    (COLLEGE OF VETERINARY AND ANIMAL SCIENCES-MANNUTHY,THRISSUR, 2011) KIRANA SASIDHARAN; K.A.Bindu
    Estrogen receptor gene and pituitary specific transcription factor 1 gene was studied as candidates for genetic markers of milk production traits. The study was aimed to reveal the known variants at ESR/TspRI and PIT/HinfI locus in Vechur and crossbred cattle of Kerala and to analyze their association with milk production. The analysis of ESR/TspRI polymorphism revealed existence of three digestion patterns (CC, CA and AA) in Vechur and crossbred cattle. The A/C allele frequencies were 0.43/0.57 and 0.53/0.47, respectively in Vechur and crossbred cattle. The genotypes of ESR/TspRI polymorphism were distributed according to Hardy-Weinberg equilibrium with frequencies 0.33(CC), 0.47 (AC) and 0.20 (AA) in Vechur and 0.24 (CC), 0.46 (AC) and 0.30 (AA) in crossbred cattle under study. PCR-RFLP analysis of 451 bp fragment of PIT-1 gene revealed the predominance of BB genotype and B allele with a mean frequency of 0.57 and 0.75, respectively. AB genotype was the next most frequent with a mean frequency of 0.39. Only six individuals out of the 130 screened were observed to have AA genotype. Allele frequency at PIT/HinfI was the same in Vechur and its crossbred with Bos Taurus. Association studies revealed the significant influence of PIT/HinfI polymorphism on milk production traits. The A allele bearing Vechur and crossbred cattle showed an increase in 305 day milk yield and average daily milk yield. Lactation length in Vechur was significantly high for AA and AB genotype. ESR/TspRI showed no significant difference in milk yield. The selection of Vechur and crossbred cattle for A allele of PIT/HinfI polymorphism can significantly improve milk yield. This study validates the utility of PCR-RFLP as efficient means of exploring the distribution pattern of known variants at important candidate genes in the cattle population of Kerala for future breeding plans.
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    ThesisItemOpen Access
    GENETIC VARIABILITY IN THE PROLACTIN RECEPTOR AND FOLLICLE STIMULATING HORMONE–β GENES OF PIG
    (COLLEGE OF VETERINARY AND ANIMAL SCIENCES-MANNUTHY,THRISSUR, 2011) DEEPA.C.N; Thirupathy Venkatachalapathy, R.
    The aim of this study was to investigate the polymorphism of the Prolactin receptor (PRLR) and follicle stimulating hormone (FSH) beta genes present in different pig breeds and its influence on reproductive traits. Reproductive traits investigated were: Litter size at birth (LSB), Litter weight at birth (LWB), litter size at viii weaning (LSW) and litter weight at birth (LWW). Genomic DNA was isolated from blood of 63 Large White Yorkshire (LWY), 25 Duroc and 32 Desi× large white Yorkshire crossbred pigs. The PCR-RFLP method was used to detect polymorphism in PRLR and FSH beta genes. Alu I polymorphism was found in 10th exon region of PRLR with two different alleles. The frequencies of PRLR allele A were 0.52 in the LWY, 0.64 in Duroc and 0.60 in the crossbreds. The allelic frequencies for PRLR- B allele were 0.48, 0.36, 0.40 in LWY, Duroc and crossbreds respectively. Hae III polymorphisms were detected in FSH beta first intron region. Two genotypes were found leading to allele frequencies of 0.87, 0.88. 0.83 for A and 0.13 ,0.12 ,0.17 for B allele in LWY, Duroc and crossbreds respectively. FSH beta fragment covering first exon and partial intronic region showed no polymorphism with Taq I in the tested sows of the LWY, Duroc and crossbreds. The association analysis between the genotypes and the reproductive traits were estimated using Least square mixed model method. PRLR/Alu I genotype showed significant association with litter size at weaning. The AB genotype had higher LSW in all breeds. Sows with the PRLR -BB genotype had the largest litter size at birth , but the difference was statistically not significant. No significant associations of FSH beta alleles with the traits were observed. Analysis were also done to estimate the effect of sire, age, breed and season of farrowing and showed that only age has significant effect on litter traits in the population studied. Sows at an age of 1.5-2 years had higher litter size and litter weight at weaning. Additional data is needed to confirm the significant effects seen in these three lines. However, the effects observed to date suggest that PRLR/Alu I polymorphism was associated with litter size at weaning and hence could be useful as a marker for litter size in selection programs.
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    ThesisItemOpen Access
    Molecular characterisation of the growth hormone gene of Vechur cattle
    (COLLEGE OF VETERINARY AND ANIMAL SCIENCES-MANNUTHY,THRISSUR, 2011) Archana.S; T.V. Aravindakshan
    The study was undertaken with the objective of molecular characterization of the growth hormone (GH) gene of Vechur cattle. It is a 22,000 dalton single-chain polypeptide hormone that stimulates growth, cell reproduction and regeneration in humans and other animals. The production of growth hormone is tissue specific and occurs predominantly within a specialized subset of cells in the anterior pituitary gland. The genomic DNA was isolated from blood samples and a 1895 bp fragment of the entire transcriptional unit of the GH was amplified by PCR using synthetic oligonucleotide primer pairs designed based on the bovine sequence (Accession # M57764; Gordon et. al., 1983) downloaded from the National Centre for Biotechnology Information (NCBI) site . The gel purified PCR product was ligated into the pGEM®-T Easy cloning vector and was transformed by giving heat shock to competent E. coli cells prepared by CaCl2 treatment. The recombinant clones among the transformed cells were identified by Blue–White Screening and the recombinant plasmid carrying the insert gene was isolated from the white clones by a modified SDS-alkaline lysis method. The ~1.9 kb GH gene insert in the vector was sequenced by the dideoxynucleotide sequencing method with primer walking using an automated DNA sequencer. The nucleotide sequence showed 99 per cent homology with Bos taurus Bos indicus ,Yak and Gayal The exon-intron boundaries in the Vechur occurred at the codons of the amino acid residues, Ala-4 (intron 1), Glu-57 (intron 2), Lys-96 (intron 3) and Met150 (intron 4). All these four residues were conserved in Bos taurus and Bos grunniens as well as in Giraffa camelopardalis. The positions of the exon-intron boundaries are also conserved in various species as evidenced from similar sizes of the exons. This strict homology in the sites of insertion of introns suggests that the exon-intron organization of these genes was established before the divergence of these species. The Vechur cattle GH gene has an open reading frame (ORF) of 654 nucleotides encoding a signal peptide of 26 amino acid residues and a mature protein of 191 amino acid residues. The 191 amino acid residues of the mature polypeptide chain was found to be 100 per cent homologous to that of Bos taurus (Accession #M57764.1). The mature amino acid sequence of growth hormone of Vechur cattle Starts with Ala at NH2- terminal and Phe at COOH-terminal. The predicted secondary structure showed that the larger α-helical lobe was formed by four sections of the polypeptide chain (residues 1-35,66-86, 111-129 and 149-191) while the smaller lobe, which encompassed a small anti-parallel beta-sheet and a small irregular structure formed the remaining structure of the polypeptide chain. The predicted 3D protein structure obtained from the Swiss Model Server and EsyPred3D and the 3D structure of human GH protein revealed that the α-helices, β-sheets and random coils in the predicted protein are similar to that of the human GH structure. This work seems to be the first of its kind in establishing the DNA and protein sequence data and structural organization of growth hormone gene in Vechur cattle. The analysis of sequence data revealed that there was high similarity of the protein at the amino acid sequence level (100 per cent identity) with Bos taurus and Bos indicus. The signal peptide and intron sequences seem to be evolved more rapidly as there was less homology as compared to exonic sequences. Overall, the nucleotide sequence of Vechur GH gene was found to be similar (99 per cent identity) to that of Bos taurus and other Bos indicus. The regulatory sequences and the amino acid sequence shows 100 per cent identity with that of Bos taurus and other Bos indicus. The study reveals that the dwarf character of this breed is not due to any change in the nucleotide sequence of GH gene. The protein sequence analysis data denoted the slow basal rate of evolution shown by mammalian GHs and also showed that the GH gene is strongly conserved among species. Signal peptide sequences seems to be evolved more rapidly as there was less homology among them as compared to mature GH sequences and there was no correlation between the rates of evolution of these two sequences.