Cloning And Sequence Of Goat ACTRA Gene And Investigation Expression By Real-time Quantitative Assays

Activin is a member of the transforming growth factor(TGF)βsuperfamily of extracellular signaling proteins. Themembers of this family play a critical role during embryogenesis and inmaintaining tissue homeo-stasis in adult life. Activins signal through cell surface serine-threonine kinase receptors that include typeⅡligand-binding receptors and typeⅠsignaling receptors. Once the ligands are bound, a typeⅡreceptor will heterodimerize and phosphorylates a typeⅠreceptor. This will activate the typeⅠreceptor, which leads to the phosphorylation of specific cytoplasmic Smad proteins. These Smads translocate to the nucleus, where they control the transcription of target genes.We used three species of goats:Dazu goat,Nanjiang goat and Saanen dairy goat in this research. nutrient-balanced, and healthy female goats at the age of 2-3, weighing 25-49kg were used, and each species has three repeats. The goats were under observe for around 30 days before testing after their first natural estrus。There has not release Capra hircus ACTR cDNA sequence in public, so far. Based on the Ovis aries and Bos taurus ACT cDNA released at the GenBank,the special primers of goat has been designed. The full-length cDNA of goat was amplificated by the RT-PCR technique using the total RNA isolated from goat ovary as template. Then its potential amino acid sequence was deduced and its protein structure was analysed.To evaluate the expression of activin receptor (ActR)on different tissues, and to investigate the regulatory effects of ACTR on different reproduction rate. In this research Dazu black goat and Nanjiang goat with high productivity and Sanen dairy goat with low productivity were used as experiment object. The concentrations of ACTR I and ACTRHin different tissues were measured by Real-time fluorescence quantitative PCR (FQ-PCR) in order to research the secretion rule about ACTR during proestrum. The main results are as following:1.The full-length cDNA of goat ACTR A gene was obtained from goat ovary as template (GenBank accession number:HQ245710,HQ701674).ACTRⅡA length was 1120 nucletides.The open reading frame(ORF) has 1088 nucletides which encoded a polypeptide of 342 amino acids; ACTRⅠA length was 1311 nucletides.The open reading frame(ORF) has 1256 nucletides which encoded a polypeptide of 418 amino acids.2.According to the Blast result in GenBank database, the cDNA nucleotides share 90% homology with different goats. Comparison of amino acids sequence of goat ACTR IIA with those of human(Homo sapiens), cattle (Bos Taurus),sheep(Ovis aries), mice (Mus musculus), the similarity were 95%,98%,99%,93%, ACTRⅠA with those of sheep(Ovis aries), cattle (Bos Taurus), orangutans (Pongo abelii),dog(Canis),human(Homo sapiens), mice (Mus musculus), the similarity were 98%,98%,98%,93%,92%,88%,93%,respectively.It showed that ACTR A gene keep highly conservative in the evolutionary process.3.The expression of ACTR A gene in different tissues was analyzed with Real-time PCR,the results showed that the ACTR A gene was expressed in spleen, lung, kidney, liver, heart, ovary and pituitary, the secretion of ACTRⅠshowed a positive correlation with ACTR II(P＜0.01); ACTRⅠand ACTRⅡin kidney is highest,and in heart, lungs is lowest. The expression level of ACTR A gene in same varieties goat with different tissues were different.The expression level of ACTR IA gene in Saanen Dairy goat were significantly higher than the Nanjiang Yellow goat and Dazu Black goat(P＜1 0.01). No significant difference was found between Nanjiang Yellow goat and Dazu Black goat about the expression level of ACTRⅡA gene (P>0.05)4.TypeⅡreceptor-mediated transmission of information in high fertility goats. It is advantageous to control typeⅡreceptor for increase the number of eggs produced.