猪骨骼肌单根肌纤维的分离与鉴定方法

发布时间：2018-06-28 06:01

  本文选题：猪 + 单根肌纤维　； 参考：《农业生物技术学报》2017年10期

【摘要】：肌纤维是构成肌肉组织的基本结构单位,肉质主要受到肌纤维类型及其组成的影响。为丰富骨骼肌生长发育的体外研究模型,本研究选取刚出生的大白猪(Sus scrofa)后肢肌肉,利用胶原酶消化法分离出单根肌纤维,并采用RT-PCR和免疫荧光染色鉴定不同肌纤维类型。对Ⅰ型胶原酶分离的单根肌纤维进行高倍镜镜检,肌纤维表面光滑并且有卵圆形凸起,其为前体骨骼肌卫星细胞,说明肌纤维具有良好活性并且其RNA未降解,可进行后续RNA微量提取实验。逆转录PCR(reverse transcription PCR,RT-PCR)结果表明,通过胶原酶消化法分离趾长伸肌的8根肌纤维为单根肌纤维,主要类型为快速酵解型(Ⅱb);并且通过慢速氧化型(Ⅰ)抗体对单根肌纤维进行免疫荧光染色验证RT-PCR结果的可靠性。本研究建立了一种简单、低成本的猪骨骼肌单根肌纤维的分离与鉴定方法,为进一步研究成熟骨骼肌细胞发育过程及其生物学功能提供了体外模型。
[Abstract]:Muscle fiber is the basic structure unit of muscle tissue. Meat quality is mainly affected by the type and composition of muscle fiber. In order to enrich the model of skeletal muscle growth and development in vitro, single muscle fiber was isolated from hind limb muscle of newly born large white pig (Sus scrofa) by collagenase digestion, and different types of muscle fibers were identified by RT-PCR and immunofluorescence staining. The single muscle fiber isolated by type I collagenase was examined by high power microscope. The surface of muscle fiber was smooth and oval protuberance. It was a satellite cell of precursor skeletal muscle, which indicated that the muscle fiber had good activity and its RNA was not degraded. The microextraction experiment of RNA can be carried out. The results of reverse transcription-PCR (reverse transcription PCR showed that 8 muscle fibers of extensor digitorum longus were separated into single muscle fibers by collagenase digestion. The main type was rapid fermentative type (鈪，

本文编号：2077008