Genetic variants of BMP15 and GDF9 genes and their association with litter size in Tibetan sheep

Introduction Bone morphogenetic protein 15 ( BMP15 ) and growth differentiation factor 9 ( GDF9 ) are oocyte-secreted members of the transforming growth factor-β superfamily involved in follicular development,ovulation, and reproductive performance. This study examined their partial sequence characteristics,tissue-expression profiles, and associations of selected single-nucleotide polymorphisms (SNPs) with litter size in Tibetan sheep. Methods Gene cloning, bioinformatic analyses, quantitative real-time PCR, and improved multiplex ligase detection reaction (iMLDR) genotyping were performed. Six ewes were used for expression profiling across 12 tissues, and 200 ewes for SNP genotyping and association analyses. Results The predicted open reading frames within the cloned BMP15 and GDF9 fragments were 468 and 405 bp, encoding 155 and 134 amino acids, respectively, and were therefore considered partial coding fragments. Their predicted amino acid sequences showed the highest similarity to Ovis aries reference sequences. The BMP15 fragment showed slight hydrophobicity, whereas the GDF9 fragment was hydrophilic overall. One transmembrane-like segment was predicted at the N-terminus of the BMP15 fragment, whereas none was detected in the GDF9 fragment. A Sec-type signal peptide was predicted in the GDF9 fragment. Exploratory qPCR analysis detected both transcripts in all tissues. BMP15 transcript abundance was highest in the oviduct, with a relatively high signal in the liver, whereas GDF9 expression was highest in the ovary. VEP classified the four SNPs as two synonymous and two missense variants, and SIFT predicted both missense substitutions to be tolerated. None of the four SNPs was significantly associated with litter size ( P > 0.05). Discussion The extra-ovarian BMP15 signals should be regarded as preliminary observations. Overall, these findings provide baseline data for further investigation of BMP15 and GDF9 in Tibetan sheep reproduction and for future identification and validation of reproduction-related functional variants.