ARMS-PCR method for detecting multiple NPM1 mutations / 中国实验血液学杂志

ABSTRACT

This study was aimed to establish a simple, sensitive detection method for multiple NPM1 mutations, so as to reduce the omission ratio of NMP1 mutant detection. Recombinant plasmids containing wide-type NPM1 and the most common mutations (A, B, C, D) were constructed as the detection objects. The ARMS-PCR for detecting multiple NPM1 mutations was established through designing a pair of specific primers whose 3' end base matched with four mutants (A,B,C,D), but did not matched with wild type NPM1 according to the different base sequence of NPM1 mutants. The feasibility of the ARMS-PCR method was evaluated by assessing the detection range and the sensitivity and comparing with direct sequencing. The results showed that the recombinant plasmids were constructed successfully by restriction analysis and DNA sequencing. The four mutants but not wild type NPM1 were detected by using ARMS-PCR, the detection range of the method was 10(3) copies/ml -10(9) copies/ml and the sensitivity was 0.01%, while the direct sequencing method could not detect the mutations if mutation was less than 10%. It is concluded that the high sensitive ARMS-PCR is established for detecting the four mutations of NPM1 and more than 95% mutants can be detected by this method, providing a new detection method for clinical NPM1 gene mutant.