“Genetic polymorphism of HSP70, LHR, LEPR genes and selective minerals status in postpartum anestrous buffaloes
Abstract
The present study was planned to obtain the functional information on genetic polymorphism
of Leptin receptor gene (LEPR), Luteinizing hormone gene (LHR) and Heat shock protein genes
(Hsp70), status of anti-oxidants, minerals and hemogram. To fulfill the present objectives twenty
postpartum anestrous (PPA) and twenty normal cyclic (NC) Murrah buffaloes were selected on the
basis of their reproductive history and current status of reproductive organs. The animals were
maintained as per the standard feeding and management. Venous blood sample was collected from
each experimental animal for DNA isolation, plasma and hemolysate separation. The 275 bp fragment
of the LHR gene, 413 bp fragment of the LEPR gene and 513bp fragment of the HSP70 gene present in
the genomic DNA of Murrah breed was amplified by using the primers pair. These partially amplified
segment were then subjected to restriction enzyme digestion and sequencing. Plasma and hemolysates
stored at -20°C were used for estimation of antioxidants status, Protein and Minerals. Blood
hematology was done with automatic hematology analyser (Vetscam 5HM, ABaxis Ltd.). The RE
analysis by AluI and MobII of partially amplified LEPR gene, by HphI of partially amplified LHR
gene and by HphI and MobII of partially amplified Hsp70 gene showed only one band pattern in both
groups animals under the study, indicates no polymorphism at these RE sites in the population studied.
After sequencing and their alignment, Single nucleotide polymorphism (SNP) was detected at 231,
247 and 347 in aligned sequence of LEPR gene and at 243 nucleotide position of LHR gene. Statistical
analysis by chi square test did not reveal any significant association of these polymorphism in relation
to PPA. After sequencing and their alignment, polymorphism at seven nucleotide positions were found
to be associated with nucleotide variations across the various samples studied (192, 256,251,258, 367,
438 and 512). Out of these six variations were consider for further analysis as one variation had have
very little representation in the samples. On applying Chi-square test, five out of six nucleotide
positions (192, 256, 258, 367 and 438) revealed no significant association between these nucleotide
variations and the health status of animal i.e. PPA and NC animals. However, a significant (P<0.05)
association was found with relation to nucleotide position 251 and, it can be inferred that nucleotide C
at position 251 is associated with postpartum anestrous condition. Since this nucleotide position is not
included in the recognition sequence of any RE, we cannot differentiate between normal and PPA
animals on the basis of RE analysis.
Description
Keywords
Postpartum anestrous, LEPR gene, LHR gene, Hsp70 gene, Antioxidants, Minerals, Vitamins