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《Chemical Journal of Chinese Universities》 2007-01
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Determineation of DNA Cleavage Site Sequence Using Fluorescence Labeled Primer and DNA Sequencer

ZHENG Wei-Juan~(1),CHEN Yuan~(1),SHAO Ying~(2),TANG Zhong-Hua~(3),GUO Zi-Jian~(2),HUA Zi-Chun~(1*)(1.State Key Laboratory of Pharmaceutical Biotechnology,2.State Key Laboratory of Coordination Chemistry,Nanjing University,Nanjing 210093,China;3.Shanghai Shenergy Biocolor Bioscience & Technology Company,Shanghai 200233,China)  
Determining the base sequence of DNA broken site is quite crucial for the study on the cleavage site specificity and mechanism of various natural or synthetic DNA cleavage regents,and on developing novel therapeutic drugs targeting at DNA.The most frequently used method depending on chemical reactions of the Maxam-Gilbert procedure,and the late arising methods used by Rui Ren et al.which were based on Sanger's DNA sequencing strategy,all had some deficiencies,either the pollution of radioactive materials,or really complicated and difficult to operate.In the present paper,a new method for DNA cleavage site sequence determination was developed.The fluorescence FAM-labeled primer was annealed to the DNA fragments,which has been cleaved by restriction enzymes or other regents,and extended along the template sequence.The products then loaded onto the polyacrylamide electrophoresis gel of ABI 377 DNA Sequencer.Data was collected and analyzed by using ABI PRISM Data Collection Software and ABI PRISM Sequencing Analysis Software.It is proved to be a credible and simple new approach to determine the base sequence of DNA broken sites.
【Fund】: 教育部优秀青年教师奖励基金(批准号:1999028418 20030284040);; 国家自然科学基金(批准号:30330530 20373026 30425009)资助
【CateGory Index】: Q78
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