Cochet M, Perrin F, Gannon F, Krust A, Chambon P, McKnight G S, Lee D C, Mayo K E, Palmiter R
Nucleic Acids Res. 1979 Jun 11;6(7):2435-52. doi: 10.1093/nar/6.7.2435.
Chicken conalbumin double-stranded cDNA (con-dscDNA) was synthesized from a laying hen oviduct mRNA preparation enriched for conalbumin mRNA (con-mRNA). The dscDNA was inserted by blunt-end ligation into the Sal I site of plasmid pBR322 which had been repaired with DNA polymerase I to create Taq I sites on each side of the inserted fragment. After bacterial transformation, one hybrid recombinant, pBR322-con1, which contains the largest inserted dscDNA (about 2350 bp) was shown to hybridize specifically to the RNA which is translated into conalbumin. Electron microscopic examination of hybrid molecules between con-mRNA and pBR322-con1 DNA indicate that the inserted con-dscDNA is an almost full-length double-stranded transcript of conalbumin mRNA.
从富含卵清蛋白mRNA(con-mRNA)的产蛋母鸡输卵管mRNA制备物中合成了鸡卵清蛋白双链cDNA(con-dscDNA)。通过平端连接将双链cDNA插入质粒pBR322的Sal I位点,该质粒已用DNA聚合酶I修复,以便在插入片段的两侧产生Taq I位点。细菌转化后,一个杂交重组体pBR322-con1被证明能与被翻译成卵清蛋白的RNA特异性杂交,该重组体包含最大的插入双链cDNA(约2350 bp)。对con-mRNA与pBR322-con1 DNA之间的杂交分子进行电子显微镜检查表明,插入的con-dscDNA是卵清蛋白mRNA的几乎全长的双链转录本。