Cloning and Testis Expression Difference Analysis of Coding Region Sequence of CRISP2 Gene in Sterile and Fertile Boar of Banna Mini-pig Inbred Line

CRISP2 gene is an important candidate gene that is linked with animal asthenozoospermia. In order to analyze the relation between CRISP2 and sperm motility and the regulation and function of CRISP2 provide basic information, CRISP2 cDNA including the coding sequence was obtained by RT-PCR with the RNA isolated from sterile and fertile boar of the Banna minipig inbred line (BMI) and submitted to GenBank with the access number KP780059, the result showing no difference in sequences between the fertile and sterile BMI boars. Bioinformatics analysis showed that the CRISP2 gene encode a protein of 244 amino acids with a predicted molecular weight of 27.05 ku and the isoelectric point of 6.76. Protein structure analysis showed that CRISP2 protein contains two conserved domains and a signal peptide. The protein without transmembrane regions had a leucine-rich nuclear export signals, its N-terminus and C-terminus are hydrophobic. The probability of secreting into periplasmic is 94.1%. Phylogenetic results indicated that the pig has the closest relationship with cattle and sheep. We tested the CRISP2 mRNA expression in 14 tissues from sterile and fertile boar of the BMI by qPCR, the result showed that the gene was specific expressed in testis, the expression in sterile boar was significantly lower than in the fertile boar (P 0.05), the reduced expression of CRISP2 may lead to the decreased sperm motility. The result of this study is a useful foundation for further insight into the role of this gene in pig spermatogenesis.