采用TA克隆技术检测红细胞增多症患者中JAK2外显子12突变的低等位基因负担

Detection of low allele burden of JAK2 exon 12 mutations using TA-cloning in patients with erythrocytosis.

作者信息

Ohyashiki Junko H, Hisatomi Hisashi, Shimizu Syoko, Sugaya Maki, Ohyashiki Kazuma

机构信息

Intractable Disease Therapeutic Research Center, Tokyo Medical University, 6-7-1 Nishishinjuku, Shinjuku-ku, Tokyo 160-0023, Japan.

出版信息

Jpn J Clin Oncol. 2009 Aug;39(8):509-13. doi: 10.1093/jjco/hyp048. Epub 2009 Jun 2.

DOI:10.1093/jjco/hyp048

PMID:19491085

Abstract

OBJECTIVE

Polycythemia vera (PV) is a clonal myeloproliferative neoplasia associated with the activation of the Janus-activating kinase 2 (JAK2) mutation. The aim of this study is to identify clonal expansion of exon 12 mutations.

METHODS

We performed DNA sequencing of the JAK2 exon 12 after TA-cloning in JAK2-V617F-negative and JAK2-V617F-positive PV patients.

RESULTS AND CONCLUSIONS

We found clonal mutations (i.e. H538-K539delinsL and D544G) in 3 of 7 JAK2-V617F-negative PV patients, however, unlike JAK2-V617F, allele burden of JAK2 exon 12 mutation was low. Since allele-specific PCR is able to amplify only the limited region which contains known mutations with gain-of-function, we need to clarify the biological implications of unknown single nucleotide substitution of the JAK2 exon 12 with low clonal burden in erythrocytosis patients.

摘要

目的

真性红细胞增多症（PV）是一种与Janus激活激酶2（JAK2）突变激活相关的克隆性骨髓增殖性肿瘤。本研究的目的是鉴定12号外显子突变的克隆性扩增。

方法

我们对JAK2-V617F阴性和JAK2-V617F阳性PV患者进行TA克隆后，对JAK2 12号外显子进行DNA测序。

结果与结论

我们在7例JAK2-V617F阴性PV患者中的3例中发现了克隆性突变（即H538-K539delinsL和D544G），然而，与JAK2-V617F不同，JAK2 12号外显子突变的等位基因负担较低。由于等位基因特异性PCR仅能扩增包含具有功能获得性的已知突变的有限区域，我们需要阐明红细胞增多症患者中JAK2 12号外显子低克隆负担的未知单核苷酸替代的生物学意义。