摘要
作者采用聚合酶链反应(PCR)成功地从含人巨细胞病毒B基因的重组质粒pBH_2DNA中扩增及分离了B基因编码区段及其产物糖蛋白52kd抗原编码区段。0.2μg的模板DNA经过30次循环,目的基因片段的扩增量达到10μg,足以用于酶谱分析及克隆的构建。
A nucleic acid amplifi-cation procedure, the polymerase chainreaction (PCR), has been used to amp-lify the human cytomegalovirus (HCMV)B gene code region, and its glycoprotein52 kd atigentic domain. The primersused in the PCR assay were based on aconserved region of the HCMV B genesequence. By using primer 1 (5′-AAAGAATTCATGGAATCCAGGATCT G-3′up-stream nucleotides 157 to 2877), primer 2(5′-AAAGAATTCATGAACGTGAAGGA-ATCG-3′,upstream nucleotides 1846 to2877), and primer 4 (5′-ATAAAGCTTAATCAGACGTTCTCTTCTTC-3′, downstreamnucleotides 157 to 2877 and 1846 to 2877) ,the HCMV B gene code region sequenceand its glycoprotein 52 kd antigentic do-main sequence were amplified from therecombintal plasmid pBH1 DNA contain-ing the HCMV B gene. Amplificationcycles consisted of denaturation at 92℃for 1 min, annaling at 55℃ for 1 min,and extension at 72℃ for 1.5 min. Thecycles were carried out 30 times in threedifferent water baths, respectively. Afterthe last extension, 10 μl of each reactionmixture was removed and subjected toelectrophoresis on 1.2% agarose gel. Ten μgof PCR products were obtained from0.2μg of template DNA after the 30times cycles, and were enough for beingused in the restriction site analysis andthe construction of clones.
出处
《华西医科大学学报》
CAS
CSCD
1991年第2期117-119,共3页
Journal of West China University of Medical Sciences
基金
高等学校博士学科点专项基金
关键词
聚合酶链反应
人巨细胞病毒
B基因
Polymerase
chain reaction
Human cytomegalovirus
B gene
coderegion
Glycoprotein
52kd antigentic domain
