摘要
本文报道了用人工合成的28聚寡核苷酸为“引物”,以强毒株新城疫 病毒(NDV)基因组PNA为模板,在逆转录酶作用下成功地合成了NDV- cDNA。后者经琼脂糖凝胶电泳观察,其大小约为0.5～14kb。从凝胶上收集 大于1.0kb以上的cDNA,用Hpa Ⅰ对其进行限制性酶切,然后将酶切片段克隆到pBR322的ScaI位点上,得到的克隆产物即为构建的NDV-cDNA文库。经用少量文库转化E.Coli HB101,得到氨苄青霉素敏感而四环素抗性的转化菌落(Amp基因因外源DNA的插入而失活)。从随机挑出的5个转化菌株中提取重组质粒,经琼脂糖凝胶电泳检测,其插入片段的大小分别为0.8～3.0kb不等。说明所构建的NDV-cDNA文库有效。
It was reported that newcastle disease virus (NDV) cDNA was succesfully synthesized using synthetic 28 oligonucleotide as primer and using genome UNA of NDV virulent strain as template with reverse transcriptase. The size of NDV-cDNA rangedO.S to 14 kb by agarose gel electrophoresls. The cDNAs above 1.0 kb were collected from the gel plate and were cut with restriction enzyme Hpa I , and then the enzyme-cut fragments were cloned at a Sea I site of pBR322 so that the NDV-cDNA library was obtained.
出处
《中国兽医科技》
CSCD
1993年第2期3-5,共3页
Chinese Journal of Veterinary Science and Technology
