Related Experiment Videos
[The nucleic acid analysis by DNA chip technique based on nuclease S1 protection]
1Department of Biological Science and Technology, School of Life Science and Biotechnology, Shanghai Jiaotong University, Shanghai 200240, China. elzeng@263.net
Objective:
Establishing a method for quantitative analysis of nucleic acid by DNA chips.
Methods:
The modified oligonucleotides with ribose at 3'-end was chemically synthesized. The 5'-end was labeled by radioisotope 32P with kinase catalyzed reactions. Such oligonucleotides were converted into di-aldehyde at 3'-end by oxidization with NaIO4, and then were spotted on glass slide with the amino group modified surface. After reduced with NaBH4, the oligonucleotides were attached strongly. The DNA chips prepared with this method were hybridized with nucleic acids existed in the solution and then digested with nuclease S1.
Results:
When they were paired with the nucleic acids in the solution perfectly, the oligonucleotides on the chip were not cleaved by nuclease S1. Otherwise, the oligonucleotides on chip were cleaved. The protection efficiencies appeared proportional to the perfect paired nucleic acids in the solution when the content of target nucleic acids were less than the spots on the slides.
Conclusions:
The method was developed for both qualitative and quantitative analysis of nucleic acid. As it was not required to label the samples with radioisotope or fluorescence, it might be a practical choice for clinical tests.