[1]韩西群,齐宗利,贺莉,等.降落式PCR和SSCP检测淋巴细胞白血病单克隆T细胞受体γ基因重排[J].南方医科大学学报,2004,(02):188-191.
 HAN Xi-qun,QI Zong-li,HE Li,et al.Touch-down PCR and single-strand conformation polymorphism for detecting clonal T cell receptor γ gene rearrangement in lymphoid leukemia[J].Journal of Southern Medical University,2004,(02):188-191.
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降落式PCR和SSCP检测淋巴细胞白血病单克隆T细胞受体γ基因重排()
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《南方医科大学学报》[ISSN:1673-4254/CN:44-1627/R]

卷:
期数:
2004年02期
页码:
188-191
栏目:
出版日期:
2004-02-01

文章信息/Info

Title:
Touch-down PCR and single-strand conformation polymorphism for detecting clonal T cell receptor γ gene rearrangement in lymphoid leukemia
作者:
韩西群1 齐宗利1 贺莉1 陆地2 赵彤1
1. 第一军医大学病理学教研室, 广东, 广州, 510515;
2. 第一军医大学细胞生物学教研室, 广东, 广州, 510515
Author(s):
HAN Xi-qun1 QI Zong-li1 HE Li1 LU Di2 ZHAO Tong1
1. Department of Pathology, First Military Medical University, Guangzhou 510515, China;
2. Department of Cell Biology, First Military Medical University, Guangzhou 510515, China
关键词:
聚合酶链反应多态现象单链构象基因重排γ链T细胞抗原受体白血病淋巴细胞
Keywords:
polymerase chain reactionpolymorphism single-stranded conformationalgene rearrangement gamma-chain T-cell antigen receptorleukemia lymphocytic
分类号:
R394.2;R733.7
摘要:
目的 以降落式PCR和单链构型多态性(SSCP)方法检测淋巴细胞白血病单克隆T细胞受体(TCR)γ基因重排,探讨其在淋巴细胞白血病诊断中的价值。方法 盐析法提取淋巴细胞白血病患者外周血白细胞DNA,TCR-γ基因重排通用引物和降落式PCR扩增基因重排。分别采用琼脂糖凝胶电泳、SSCP和DNA直接测序方法检测PCR扩增产物。阳性细胞系DNA模板与反应增生性淋巴组织DNA按不同比例混合后扩增,检测降落式PCR的灵敏度。结果 琼脂糖电泳中,18例T淋巴细胞白血病中有15例、4例B淋巴细胞白血病中有2例为阳性,阳性标本经SSCP进一步分析,呈不连续条带。DNA直接测序证实扩增产物为TCR-γ基因重排。阳性对照模板占1%以上时可得到阳性扩增结果。结论 TCR-γ基因重排通用引物结合降落式PCR可有效扩增T淋巴细胞白血病基因重排,可用于T淋巴细胞白血病的辅助诊断。
Abstract:
Objective To explore the value of detecting clonal T cell receptor γ (TCR-γ) gene rearrangement with touch-down PCR and single-strand conformational polymorphism analysis (SSCP) in the diagnosis of lymphoid leukemia. Methods The DNA of peripheral blood leucocytes from lymphoid leukemia patients were extracted for amplification of the TCR-γ gene rearrangement with the consensus primers and touch-down PCR. The PCR products were analyzed by agarose gel electrophoresis, direct DNA sequencing and SSCP analysis. The positive control cell line DNA was mixed in different proportions with the DNA extracted from reactive lymphoid tissue to test the sensitivity of the touch-down PCR. Results Fifteen of 18 T lymphoid leukemia and 2 of the 4 B lymphoid leukemia patients were identified to be positive by agarose electrophoresis. The positive PCR products were further analyzed by SSCP analysis, which showed discrete bands. Direct DNA sequencing confirmed the clones to be TCR-γ gene rearrangement, and the sensitivity of touch-down PCR was 1%. Conclusion Consensus primers for studying TCR-γ gene rearrangement in combination with touch-down PCR can effectively amplify the clonal TCR-γ gene rearrangement in T lymphoid leukemia.

参考文献/References:

[1] Macintyre EA, Delabesse E. Molecular approaches to the diagnosis and evaluation of lymphoid malignancies[J]. Semin Hematol, 1999,36(4): 373-89.
[2] Gebhard S, Benhattar J, Bricod C, et al. Polymerase chain reaction in the diagnosis of T-cell lymphoma in paraffin-embedded bone marrow biopsies: a comparative study[J]. Histopathology, 2001, 38(1): 37-44.
[3] Kneba M, Bolz I, Linke B, et al. Characterization of clone-specific rearrangement T-cell receptor gamma-chain genes in lymphomas and leukemias by the polymerase chain reaction and DNA sequencing[J]. Blood, 1994, 84(2): 574-81.
[4] Strominger JL. Developmental biology of T cell receptors[J].Science, 1989, 26, 244(4907): 943-50.
[5] Zhu D, Kadin ME, Samoszuk M. Detection of clonal T-cell receptorgamma gene rearrangement by PCR/temporal temperature gradient gel electrophoresis[J]. Am J Clin Pathol, 2001, 116(4): 527-34.
[6] Fodinger M, Buchmayer H, Schwarzinger I, et al. Multiplex PCR for rapid detection ofT-cell receptor-gamma chain gene rearrangements in patients with lymphoproliferative diseases[J]. Br J Haematol,1996, 94(1): 136-9.
[7] Larsen HH, Masur H, Kovacs JA, et al. Development and evaluation of a quantitative, touch-down, real-time PCR assay for diagnosing Pneumocystis carinii pneumonia[J]. J Clin Microbiol, 2002, 40(2):490-4.
[8] Wack A, Montagna D, Dellabona P, et al. An improved PCR-heteroduplex method permits high-sensitivity detection of clonal expansions in complex T cell populations[J]. J Immunol Methods,1996, 196(2): 181-92.
[9] Langerak AW, Szczepanski T, van der Burg M, et al. Heteroduplex PCR analysis ofrearrangod T cell receptor genes for clonality assessment in suspect T cell proliferations[J]. Leukemia, 1997, 11 (12): 2192-9.
[10] Crisi GM, Emanuel JR, Johnson C, et al. Semireannealing, singlestranded conformational polymorphism: a novel and effective tool for the diagnosis of T-cell locality[J]. Diagn Mol Pathol, 2002, 11(2): 67-74.
[11] Trainor KJ, Brisco MJ, Wan JH, et al. Gene rearrangement in B-and T-lymphoproliferative disease detected by the polymerase chain reaction[J]. Blood, 1991, 78(1): 192-6.

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备注/Memo

备注/Memo:
收稿日期:2003-10-11。
基金项目:广东省科技计划项目(B30101)
作者简介:韩西群(1971-),男,第一军医大学在读博士研究生,医师,电话:020-61363263,E-mail:hanxiqun@fimmu.com
更新日期/Last Update: 1900-01-01