摘要
目的:构建重症肌无力(MG)患者胸腺组织抑制性消减cDNA文库,分析MG胸腺差异表达基因。方法:分别从6例正常和6例MG患者胸腺组织中分离出mRNA,逆转录合成cDNA,行抑制性消减杂交,将得到的差异表达基因进行T-A克隆,构建cDNA文库,并进行序列分析。应用实时荧光定量PCR检测20例MG患者、10例正常对照胸腺组织中GSTM3和KPNA5 mRNA的表达。结果:共获得125个阳性克隆;Blast同源性检索共得到27个差异表达基因;MG患者GSTM3、KPNA5基因表达量(0.671±0.097和0.712±0.080)高于正常对照胸腺(0.582±0.047和0.571±0.018),差异有统计学意义(tGSTM3=5.458,P<0.001;tKPNA5=2.755,P=0.010),与消减文库结果一致。结论:成功建立了MG胸腺组织抑制性消减cDNA文库。
Aim：To build suppression subtractive cDNA library of thymus from myasthenia gravis（MG） patients.Methods：The total mRNA was extracted from thymus of normal people and MG patients and synthethized to cDNA by reverse transcription.The differentially expressed genes obtained by using suppression subtractive hybridization were proceeded by T-A cloning and sequence analysis.The expressions of GSTM3 and KPNA5 in thymus of 20 MG patients and 10 normal people were detected by real-time quantitative PCR.Results：A total of 125 positive clones were obtained;27 differentially expressed genes were obtained by Blast homology search.The expression levels of GSTM3,KPNA5 in thymus tissue of MG patients（0.671±0.097,0.712±0.080）were significantly higher than those of the control group（0.582±0.047,0.571±0.018）（tGSTM3=5.458,P0.001;tKPNA5=2.755,P=0.010）,consistent with the result of subtractive library.Conclusion：The establishment of suppression subtractive cDNA library of MG thymus is successful.
出处
《郑州大学学报（医学版）》
CAS
北大核心
2012年第3期295-299,共5页
Journal of Zhengzhou University(Medical Sciences)
基金
河南省杰出青年基金资助项目0212000100
河南省科研院所专项基金资助项目0441130303
关键词
抑制性消减
重症肌无力
差异基因
CDNA文库
胸腺
suppression subtractive
myasthenia gravis
differential gene
cDNA library
thymus
