DOUBLE DIGEST RESTRICTION ASSOCIATED DNA SEQUENCING FOR IDENTIFICATION OF PROLIFICACY RELATED SINGLE NUCLEOTIDE POLYMORPHISM MARKERS IN GOATS
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Date
2018
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College of Veterinary and animal Science,Mannuthy
Abstract
Goats, commonly called as ‘poor man’s cow’ forms the basis of livelihood
for economically weaker sections of society. India has 23 well defined goat breeds,
of which Kerala has two native breeds, Malabari and Attappady Black, which vary
significantly in prolificacy. Double digest restriction associated DNA sequencing
(ddRADseq) is a modern reduced representation sequencing technique which is a
powerful and inexpensive approach for identification of Single Nucleotide
Polymorphisms (SNP) across populations. In this study ddRADseq technique was
applied to highly prolific Malabari and low prolific Attappady Black goats to
unravel the genetic variations existing between them. ddRADseq discovered 4140
SNPs and 126 INDELS in Malabari and 2192 SNPs and 59 INDELS in Attappady
Black goats of which 55 per cent variants were common between the groups. A panel
of 38 candidate prolific genes were screened and 25 SNPs were discovered in 20
genes which included one exonic variant and 18 intronic variants. Another set of 11
genes were identified based on the difference in allelic distribution of SNPs between
the two groups, which might have an effect on prolificacy. Based on in silico studies,
variant c.614-32789C>T in BMP6 was chosen for validation studies in 100 Malabari
and 50 Attappady Black goats. PCR-RFLP revealed all the three genotypes TT, TC
and CC, thus confirming the robustness of SNPs discovered by ddRADseq.
SERPIND1 of super family Serpins is reported to have a major role in placenta
formation in humans. Variant c.1259T>C identified in exon three of SERPIND1,
showed significant association with litter size with a mean value of 1.348±0.071 for
TT genotype and 2.27 ±.0.132 for TC genotype. This emphasized the relevance of
this polymorphism and usefulness of ddRADseq in identifying novel SNPs in
unexplored genes.RGS 14 gene which affects early embryonic development,
harboured a missense mutation c.1624G>A resulting in p.Pro542Ser, suggesting a
variation in protein quality. Expression study of RGS 14gene in endometrial tissues
revealed a 3.35 fold higher expression in Malabari than Attappady Black goats which
was statistically significant. Current study emphasized the relevance of ddRADseq in
identifying novel SNPs in candidate genes responsible for economic traits and
unexplored genes affecting complex traits.