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《CHINESE JOURNAL OF MICROECOLOGY》 2000-01
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A RAPID SCREEN FOR RECOMBINANT CLONES BY POLYMERASE CHAIN REACTION

Hu Wei,Xiang Hua,Zhou Yan,et al. (Basic Medical Research Center,Beijing Red Cross Chaoyang Hospital, Beijing,100020)  
Objective:To develop a polymerase chain reaction(PCR) technique for rapid screening of phenylalanine ammonia lyase(PAL) cDNA recombinant plasmid. Methods:PCR primers were designed so that one primer is complementary to the T 7 promoter of pET23b vector,and another primer complementary to the 3′ end of PAL cDNA.The recombinant colonies were transferred by using sterile pipet tip into the PCR reaction mixture directly.After PCR amplification,the PCR products were subjected to electrophoresis on 0 8% agrose gel. Results:2 positive clones with a correct direction by using the screening method were detected.The positive clones were further confirmed by DNA sequencing. Conclusions:Recombinant vectors in bacterial cells can be analyzed directly with the use of PCR. This convenient and timesaving procedure eliminates the need for preparation and purification of the plasmid.
【Fund】: 北京自然科学基金
【CateGory Index】: Q785
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