角蛋白71(Keratins 71,KRT71)属于人类Ⅱ型上皮角蛋白,是毛囊特异性的上皮角蛋白基因。相关研究表明,在其他物种中,KRT71与被毛卷曲相关,KRT71在卷曲的被毛形成过程中起着重要的作用。现眼观羊驼背部、耳部和腿部毛发的弯曲程度不同,但KRT71与其被毛的弯曲度是否相关尚不清楚。本研究利用PCR技术克隆KRT71基因全CDS区。通过实时荧光定量PCR技术检测基因的表达。免疫组化、Western印迹对KRT71蛋白在羊驼背部、耳部和腿部皮肤组织中的表达量进行分析,以探讨KRT71与羊驼毛发弯曲度的关系。结果显示,羊驼的KRT71基因的CDS区共有1 578 bp,编码525个氨基酸;免疫组化结果显示,KRT71在羊驼的背部、耳部和腿部均有表达,并且主要特异性表达于毛囊的内根鞘;实时荧光定量PCR和Western印迹结果显示,KRT71的mRNA和蛋白量均呈现背部﹥腿部﹥耳部的趋势;相较于耳部,背部和腿部的表达量较高。实验结果提示,KRT71的表达量与毛纤维的弯曲度呈正相关,KRT71可能在羊驼毛纤维的弯曲度形成过程中发挥着一定的作用。
Keratins 71( KRT71) belong to the human typeⅡepithelial keratin,as one of the hair follicle epithelium specific keratin genes. It has been shown that KRT71 associated with curly hairs. The curly coat in back,ears and leg of alpaca appeared in different degrees,but whether it related to KRT71 expression is unclear. In this study,the complete CDS region of KRT71 in 1578 bp was cloned,encoding a protein of 525 amino acids. Using real-time PCR,immunohistochemistry and Western blotting,the RNA and protein expression of KRT71 in back, ear and leg skin tissue were analyzed.Immunohistochemistry results showed that KRT71 were expressed in the back,ears and leg of alpaca.The positive staining was found in the inner root sheath of hair follicles. The expression of KRT71 mRNA and protein were also detectable in the back,leg and ear. The expression in the ear skin was lower than those in the skins from back and leg. These results suggested that KRT71 might play a role in the formation of the alpaca curly coat.