【目的】为了预测无关捐献者干细胞移植(HSCT)后GVHD的发生及程度和选择相容的干细胞捐献者,通过同时采用PCR-序列特异性引物扩增技术(sequence specific primer,PCR-SSP)和PCR-直接碱基序列分析基因分型技术(sequence based genotyping...【目的】为了预测无关捐献者干细胞移植(HSCT)后GVHD的发生及程度和选择相容的干细胞捐献者,通过同时采用PCR-序列特异性引物扩增技术(sequence specific primer,PCR-SSP)和PCR-直接碱基序列分析基因分型技术(sequence based genotyping,PCR-SBT)的方法,提高HLA基因分型的准确性和分辨水平。【方法】采用PCR-SSP和PCR-SBT分别对57份捐献者血样进行HLA-A、B和DRB1位点的高分辨等位基因分型(即识别基因的编码达到*后4位数)。【结果】PCR-SBT实验中有27份DNA的基因分析结果模棱两可,PCR-SSP实验中有10份DNA的基因分析结果模棱两可。经两种方法相互佐证实验后,57份样本均获得结果一致、清楚和精确的HLA-A、B、DRB1位点高分辨基因分型。【结论】PCR-SSP和PCR- SBT是HLA高分辨基因分析的标准方法,两种方法具有实验互补意义,若同时应用两种方法能够有效改善HLA等位基因高分辨分型的准确性和重复一致率。展开更多
Background. Mutational activation of KRAS and BRAF protooncogenes contributes to the development of many hu man cancers. Current research on gynecological cancer and specifically in cervical and endometrial cancer is ...Background. Mutational activation of KRAS and BRAF protooncogenes contributes to the development of many hu man cancers. Current research on gynecological cancer and specifically in cervical and endometrial cancer is focused on the mechanisms of their mutational activation. Objectives. In view of the paucity of data on their mutation frequency and the status of BRAF in these two types of gynecological cancer, we performed a systematic molecular study in 114 clinically and histologically well- defined malignant tumors of uterine cervix and endometrium and correlated the mutation status of KRAS and BRAF with the age at diagnosis and with tumor grade, stage or histological type. Methods. Direct sequence analysis of the PCR products of KRAS and BRAF genes was used to screen for known activating mutations. Results. In 67 cases of endometrial cancer, six KRAS mutations (8.9% ) were found, four at codon 12 (5.9% ) and two at codon 13 (2.9% ),while no mutation was detected at codon 61. Most of the mutations occurred in surgical stage I and in the endometrioid adenocarcinoma subtype. We also detected three KRAS point mutations (6.3% ) in the 47 cervical cancer samples, two at codon 12 (4.2% ) and one at codon 13 (2.1% ), while there was no mutation at codon 61. On the contrary,nomutation was identified inBRAF exon 15 for either endometrial or cervical cancer samples at position V600, which represents the most frequently mutated site of BRAF in human cancer. There was no association between KRAS mutations with either histological type, tumor grade or stage. Interestingly, however, KRAS mutation status in endometrial cancer was strongly associated with increased age at diagnosis (P < 0.001). Conclusions. Our data document (a) the absence of BRAF mutations in cervical and endometrial cancer, despite the mutation status of KRAS, (b) suggest that KRAS mutations reflect an early event in endometrial carcinogenesis and (c) imply that BRAF activation is involving alternative pathways in these two types of cancer.展开更多
文摘【目的】为了预测无关捐献者干细胞移植(HSCT)后GVHD的发生及程度和选择相容的干细胞捐献者,通过同时采用PCR-序列特异性引物扩增技术(sequence specific primer,PCR-SSP)和PCR-直接碱基序列分析基因分型技术(sequence based genotyping,PCR-SBT)的方法,提高HLA基因分型的准确性和分辨水平。【方法】采用PCR-SSP和PCR-SBT分别对57份捐献者血样进行HLA-A、B和DRB1位点的高分辨等位基因分型(即识别基因的编码达到*后4位数)。【结果】PCR-SBT实验中有27份DNA的基因分析结果模棱两可,PCR-SSP实验中有10份DNA的基因分析结果模棱两可。经两种方法相互佐证实验后,57份样本均获得结果一致、清楚和精确的HLA-A、B、DRB1位点高分辨基因分型。【结论】PCR-SSP和PCR- SBT是HLA高分辨基因分析的标准方法,两种方法具有实验互补意义,若同时应用两种方法能够有效改善HLA等位基因高分辨分型的准确性和重复一致率。
文摘Background. Mutational activation of KRAS and BRAF protooncogenes contributes to the development of many hu man cancers. Current research on gynecological cancer and specifically in cervical and endometrial cancer is focused on the mechanisms of their mutational activation. Objectives. In view of the paucity of data on their mutation frequency and the status of BRAF in these two types of gynecological cancer, we performed a systematic molecular study in 114 clinically and histologically well- defined malignant tumors of uterine cervix and endometrium and correlated the mutation status of KRAS and BRAF with the age at diagnosis and with tumor grade, stage or histological type. Methods. Direct sequence analysis of the PCR products of KRAS and BRAF genes was used to screen for known activating mutations. Results. In 67 cases of endometrial cancer, six KRAS mutations (8.9% ) were found, four at codon 12 (5.9% ) and two at codon 13 (2.9% ),while no mutation was detected at codon 61. Most of the mutations occurred in surgical stage I and in the endometrioid adenocarcinoma subtype. We also detected three KRAS point mutations (6.3% ) in the 47 cervical cancer samples, two at codon 12 (4.2% ) and one at codon 13 (2.1% ), while there was no mutation at codon 61. On the contrary,nomutation was identified inBRAF exon 15 for either endometrial or cervical cancer samples at position V600, which represents the most frequently mutated site of BRAF in human cancer. There was no association between KRAS mutations with either histological type, tumor grade or stage. Interestingly, however, KRAS mutation status in endometrial cancer was strongly associated with increased age at diagnosis (P < 0.001). Conclusions. Our data document (a) the absence of BRAF mutations in cervical and endometrial cancer, despite the mutation status of KRAS, (b) suggest that KRAS mutations reflect an early event in endometrial carcinogenesis and (c) imply that BRAF activation is involving alternative pathways in these two types of cancer.
