Detection and molecular cloning of growth differentiation factor-9 transcripts in In vitro matured buffalo oocyte complexes

The present study was carried out on buffalo ovarian follicles. From these follicles cumulus oocyte complexes were aspirated and in vitro matured in TCM-199 supplemented 10% v/v fetal bovine serum, 1μg/ml estradiol, 10μg/ml LH, 5μg/ml FSH and 1% v/v gentamycin. in vitromatured cumulus oocyte complexes were investigated for the presence of GDF -9 gene transcript using RT-PCR analysis. After the RT-PCR analysis the amplified product was purified and cloned. The cloned product was further sequenced and analyzed to study the phylogenetic relationship of GDF-9 gene of buffalo with other species. Upon RT-PCR analysis it was revealed that GDF-9 transcripts were expressed exclusively in in vitromatured cumulus oocyte complexes. Nucleotide sequence analysis of GDF-9 gene of Bubalus bubalis revealed that it is 99% similar to that of Bos taurus and clustered together with Bos taurus and Rattus norvegicus shared a separate cluster.