绵羊骨骼肌快速肌钙蛋白(TnCf)电子克隆及RT-PCR验证
摘要
Identification of sheep fast skeletal muscle troponin C (TnCƒ) gene based on its sequence conservation has two major approaches, low-stringency hybridization of cDNA libraries, and PCR method using degenerate primers. Both of the two approaches are technically difficult. In current study we have developed an in silico method to isolate the sheep TnCƒ gene and confirmed the in silico cloning by RT-PCR. After the completely sequencing of cloned cDNA, we found the sheep TnCƒ cDNA contained an open reading frame (ORF) consisting of 483 base pairs (bp) that encoded 160 amino acids (aa). No difference was observed in nucleotide composition of TnCƒ cDNA between in silico and cloned cDNA sequences, except for 4 base pairs. It suggested that in silico is an efficient method for gene cloning.