摘要
loc558117基因是trim45在斑马鱼中的同源基因,小鼠及果蝇研究结果表明该基因与血细胞发育有关.利用PCR技术扩增斑马鱼loc558117基因部分编码区,并将其插入到原核表达载体pET-28a中,经过酶切和测序鉴定后,将重组质粒(pET-28a-loc558117)转入Rosseta感受态细胞,通过IPTG诱导表达融合蛋白,his柱子亲和纯化,将纯化的融合蛋白免疫新西兰大白兔,制备多克隆抗体,并用Western Blot检测抗体.获得了loc558117原核表达重组融合蛋白及高效价的特异性兔抗loc558117多克隆抗体.为进一步研究loc558117功能提供了有力工具.
The loc558117 gene is the homologous genes of the trim45 in zebrafish, previous studies showed that it had some relations with hematopoiesis in mouse and drosophila. In this work, loc558117 gene was amplified by PCR and then cloned into pET-28a vector. After identification by restriction enzyme and sequencing, the recombinant expression plasmid containing loc558117 gene was transformed into Rosseta cells. The loc558117 fusion protein was induced by IPTG and purified protein using his-IDA sephrose, the loc558117 fusion protein was used to immuned the New Zealand white rabbits to prepare antibody. The antibody titer and specification were identified by Western blot. His fusion protein of loc558117 was obtained, and high sensitivity and specificity anti-loc558117 polyclonal antibody was gotten. The results will be helpful for functional studies of loc558117 gene in the future.
出处
《湖南师范大学自然科学学报》
CAS
北大核心
2011年第2期75-78,共4页
Journal of Natural Science of Hunan Normal University
基金
国家自然科学基金资助项目(31071999,30771170,30971105,30971663,30871340,30671053,30970425,30900851)
湖南省自然科学基金资助项目(10JJ1006)
