二步聚合酶链反应检测皮肤T细胞淋巴瘤的T细胞受体基因重排x.docx

网址： HYPERLINK  第  PAGE 4 页，共  NUMPAGES 4 页 二步聚合酶链反应检测皮肤T细胞淋巴瘤的T细胞受体基因重排 二步聚合酶链反应检测皮肤T细胞淋巴瘤的T细胞受体基因重排 中华皮肤科杂志 1998年第4期第31卷 论著 作者：丁铁钢　邱丙森　Jack Longley 单位：美国耶鲁大学医学院皮肤科(丁铁钢，Jack Longley)；上海医科大学皮肤病学研究所皮肤病理研究室(邱丙森) 关键词：聚合酶链反应；T细胞受体；基因重排；皮肤T细胞淋巴瘤 【摘要】　目的　应用二步聚合酶链反应(PCR)检测皮肤T细胞淋巴瘤(CTCL)的T细胞受体(TCR)基因重排，提高其准确性和特异性，并应用已知重排的cDNA序列，推测该病患者的特异肽链顺序，为免疫治疗提供新线索和途径。方法　先用TCR基因重排区域以外的部分为引物，做10～30个循环扩增。取得一定数量的cDNA，作为模板，再经30～40个循环扩增，获得较纯的cDNA。然后将此产物放入质粒，进一步分析DNA序列。按照分析结果，制备出特异的分子探针，对二步PCR扩增产物进行分子杂交，推测出患者的特异性肽链顺序。结果　以Vβ8为特殊引物所显示的反应带明显，用Vβ8顺序制成的分子探针杂交，示明显的特异性结合。特异性分子探针只与CTCL患者的cDNA杂交，与对照组标本不发生交叉反应。结论　二步PCR可准确地检测CTCL的TCR基因重排，其特异性强，并可在此基础上应用已知重排的cDNA序列推测该病患者的特异性肽链，为免疫治疗提供新线索和途径。 Two Step Polymerase Chain Reaction Detecting the T-Cell Receptor Gene Rearrangement in Cutaneous T-Cell LymphomaDing Tiegang*,Qiu Bingsen, Jack Longley. *Department of Dermatology, Medical College,Yale University, USA 【Abstract】　Objective　A two step polymerase chain reaction (PCR) strategy was used to amplify copy DNAs corresponding to the special region of the T-cell receptor (TCR)β-chain.Methods and Results With this technique, a dominant TCR gene rearrangement that used the β8 variable region gene segment was identified in a patient with cuteneous T cell lymphoma and its sequence determined from multiple independently generated subclonal PCR products. An oligonucleotide corresponding to thehighly variable junctional region of the clonally expanded TCR gene rearrangement was synthesized and used for further confirming studies. Finally, from the sequence data obtained, a peptide corresponding to the β-chain junctional regionsequence of the patient′s clonally expanded T-cells was generated.Conclusion Preliminarystudies using this peptide indicate that it is antigenic and may potentially beuseful in developing a patient-specific tumour vaccine. Key Words　Polymerase chain reactionT-cell receptorGenerearrangementCutaneous T-cell lymphoma T细胞在发生肿