A MutS-based protein chip for detection of DNA mutations

作者：Bi LJ; Zhou YF; Zhang XE^*; Deng JY; Zhang ZP; Xie B; Zhang CG

来源：Analytical Chemistry, 2003, 75(16): 4113-4119.

DOI：10.1021/ac020719k

This paper describes a new protein chip method for detection of single-base mismatches and unpaired bases of DNA, using a genetic fusion molecular system Trx-His(6)-Linker peptide-Strep-tagII-linker peptide-MutS (THLSLM). The THLSLM coding sequence was constructed by attaching Strep-tag 11 and mutS gene to pET32a (+) sequentially with insertion of a linker peptide coding sequence before and behind Strep-tagII gene, respectively. THLSLM was expressed in E. coli AD494 (DE3) and purified using Ni2+-chelation affinity resin. THLSLM retained both mismatch recognition activity and streptavidin binding affinity. THLSLM was then immobilized on the chip matrix coated with streptavidin through the Strep-tag II-streptavidin binding reaction. The resulting protein chip was used to detect the mismatched and unpaired mutations in the synthesized oligonucleotides, as well as a single-base mutation in rpoB gene from Mycobacterium tuberculosis, with high specificity. The method could potentially serve as a platform to develop the high-throughput technology for screening and analysis of genetic mutations.

• 出版日期2003-8-15
• 单位中国科学院武汉病毒研究所