利用寡核苷酸芯片诊断α-地中海贫血无创方法的探讨

Study of prenatal diagnosis in α-thalassemia by non-invasive method based on oligonucleotide microarray

目的通过比较地中海贫血脐血与正常脐血和外周血的全基因组表达差异,进一步探讨早期诊断地中海贫血的无创方法。方法利用含有人约22 000条Oligo DNA的人类全基因长寡核苷酸芯片,对1例α地中海贫血患儿脐血与1例正常脐血及1例外周血分别配对检测差异基因表达,并用real-time PCR验证差异基因HBA2在脐血和外周血中的差异性。采用分层聚类分析等显著性检验方法发现统计学意义上差异表达的基因。结果两组芯片结果显示,检测到的基因共4 205条,其中两者差异的基因共822条(ratio≥2或≤0.5);通过间接比较两组差异率为19.5%,即重复率为81.5%。实时定量PCR(RT-PCR)的Ct(threshold cycle)值结果亦显示HBA2在脐血和外周血中表达无差异。结论高通量的基因芯片技术能够筛选出大量的地中海贫血差异表达基因,直接与α-地中海贫血相关的HBA2蛋白在脐血和外周血中的表达无差异。预示着利用微阵列分析法对地中海贫血进行早期诊断时,可以采用无创技术制备芯片样本。

Objective To analyze differentially expressed genes between thalassemia and normal one and peripheral blood and to study the minimally invasive method for prenatal diagnosis. Methods Gene-expression profiles of 1 sample of α-thalassemia and 1 sample of normal cord blood and peripheral blood were analyzed by unsupervised Hierarchical clustering in order to get the specifically differentially expressed genes. The gene expression presence of HBA2 was verified by real-time PCR. Results A total of 4 205 genes of thalassemia were checked. Of them,822 genes were the differential genes in both groups （ratio≥2.0 or 40.05）. The differential rate was 19.5% ,i. e. , their recurrence rate was 81.5%. There were no significant differences between cord blood and peripheral blood by the value of Ct in HBA2. Conclusion These data are helpful for a better understanding of thalassemia and contribute to the development of diagnostic and therapeutic strategies. There were no significant differences between cord blood and peripheral blood in the expressions of α-thalassemia by the gene of HBA2. It is prognosticated we can use the non-invasive method for prenatal diagnosis in thalassemia by the method of microarry.