摘要
目的应用新型MGB探针的实时荧光定量聚合酶链反应(FQ-PCR)分析方法,研究mdr1 mRNA在K562细胞系和其耐药细胞系K562/A02细胞及急性白血病(AL)患儿细胞中的表达水平。方法运用Taq Man-MGB探针,以含有目的基因mdr1cDNA的质粒pHaMDR1/A为阳性模板,设立实时FQ-PCR反应体系,研究mdr1 mRNA在不同类型、不同分期AL患儿白血病细胞中的表达;并就AL患儿初治组、难治复发组、完全缓解组分别与正常对照组比较其mdr1 mRNA表达率和表达量,且在AL患儿3组间再作mdr1 mRNA表达率及其表达量比较,数据均作统计学处理。结果初治和完全缓解期AL患儿mdr1 mRNA表达阳性率和表达水平均较正常对照组升高。难治复发组AL患者白血病细胞mdr1 mRNA表达阳性率和表达水平较其他AL各组明显升高;AL完全缓解患者维持期化疗的随访观察中,部分患儿mdr1 mRNA表达量渐升高,尤其是复发AL患儿升高更显著。结论AL患儿化疗耐药的发生与mdr1 mRNA表达有较密切的联系,应用Taq Man-MGB荧光探针进行实时FQ-PCR检测人类mdr1基因在AL细胞中表达,可为AL患儿个体化化疗方案的合理制定、化疗疗效和预后判断及针对性逆转由于mdr1基因表达导致的多药耐药提供有价值的实验室依据。
Objective To study the expression of mdr1 - mRNA in K562 cell line,drug fast cell line K562/A02 and children acute leukemia （AL） ceils of different type and different periods with a fluorescent quantity polymerase chain reaction（FQ- PCR） using MGB probe. Methods A FQ- PCR system was used to detect human mdr1 gene when pHaMDR1/A was used as positive template. Isolate mononuclear cell of bone marrow and peripheral blood, then extract RNA, FQ- PCR was used to detect the expression of mdr1 after reverse transeriptase reaction. Statistical analysis of mdr1 expression rate and quantity was done between different groups. Results Among control group, different typies and periods of AL, their mdr1 - mRNA positive ratio and mdr1 - mRNA level had statistical differences. The mdr1 - mRNA positive rate and expression level in initial treatment AL group and complete remission AL group were higher than those in control group. The mdr1 - mRNA positive rate and expression level on refractory and relapse group were the highest among other AL groups. In the complete remission AL group,the expression level of mdr1 - mRNA was gradually high in some patients, especially higher when they relapse. Conclusions Taq Man - MGB fluorescent probe is used to establish a FQ - PCR system to detect human mdr1 gene. The minimum is 15 DNA copies each reaction. The above results demonstrate that the development of MDR in AL patients have more relative connection to mdr1 - mRNA,mdr1 expression can give important and valueable clue to clinical individualize chemotherapy, judgement of chemotherapeutic effect and prognosis, the research on reverse multidrug resistance.
出处
《实用儿科临床杂志》
CAS
CSCD
北大核心
2006年第21期1486-1488,共3页
Journal of Applied Clinical Pediatrics
基金
广东省医学科学技术研究基金项目资助(A2005288)
关键词
白血病
急性
多药耐药
实时荧光定量聚合酶链反应
儿童
acute leukemia
multidrug resistance
real time fluorescent quantitation polymerase chain resction
child
