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目的 检测广州地区鼻咽癌组织EB病毒LMP1基因N -末端区XhoI酶切位点的丢失 ,探讨LMP1基因变异在鼻咽癌发生发展中的作用。方法 收集中山大学肿瘤防治中心鼻咽癌新鲜活检标本 63例以及EB病毒健康携带者外周血单个核细胞 10例。采用QIAampDNAMiniKit和QIAampDNABloodMiniKit分别抽取组织和外周血单个核细胞的DNA ,应用巢式PCR扩增EB病毒LMP1基因的N -末端区 ,并用XhoI对扩增产物进行酶切。采用四色荧光末端终止法对扩增产物进行序列分析。结果 10例健康携带者外周血单个核细胞的EB病毒LMP1基因N -末端区均未见XhoI酶切位点的丢失。 63例鼻咽癌组织中有 5 0例 ( 79%)出现XhoI酶切位点的丢失 (XhoI -loss) ,4例 ( 6%)为XhoI酶切位点部分丢失 ,只有 9例 ( 14 %)未见XhoI酶切位点的丢失 (wt -XhoI)。除了XhoI酶切位点的丢失 (nt:16942 3～16942 8;GAGCTC→GATCTC)外 ,还发现 4个错义点突变。结论 广州地区EB病毒健康携带者外周血单个核细胞所携带的EB病毒LMP1基因为wt -XhoI ,而在鼻咽癌组织中主要为XhoI -loss。因此 ,我们认为EB病毒LMP1基因N -末端区XhoI酶切位点的丢失和其他的错义点突变可能是在鼻咽癌的发生发展过程中产生的。
Objective To detect the loss of XhoI restriction site in the N - terminal region of Epstein - Barr virus LMP1 gene in nasopharyngeal carcinoma of Guangzhou and to explore the role of LMP1 gene mutation in the occurrence and development of. Methods 63 fresh nasopharyngeal biopsy specimens from Sun Yat-sen University Cancer Center and 10 peripheral blood mononuclear cells from healthy carriers of Epstein-Barr virus were collected. The DNA of tissue and peripheral blood mononuclear cells were extracted by QIAampDNAMiniKit and QIAampDNABloodMiniKit. The N - terminal region of Epstein - Barr virus LMP1 gene was amplified by nested PCR and the amplified product was digested by XhoI. The amplification products were sequenced by four-color fluorescence termination. Results No XhoI restriction sites were found in the N - terminal region of Epstein - Barr virus LMP1 gene in peripheral blood mononuclear cells from 10 healthy donors. XhoI-loss was found in 50 cases (79%) of 63 cases of nasopharyngeal carcinoma, and XhoI-loss was lost in 4 cases (6%). Only 9 cases (14% No loss of XhoI restriction sites (wt-XhoI) was observed. In addition to the loss of XhoI restriction sites (nt: 16942 3 to 16942 8; GAGCTC → GATCTC), four missense point mutations were also found. Conclusion Epstein-Barr virus (LMP1) gene of Epstein-Barr virus (Epstein-Barr virus) carrying Epstein-Barr virus (EBV) healthy carriers in Guangzhou is wt-XhoI and mainly XhoI-loss in NPC tissues. Therefore, we think that the deletion of the XhoI restriction site of the N - terminal region of Epstein - Barr virus LMP1 gene and other missense point mutations may be generated during the development of NPC.