Inter Alu PCR 技术在鉴定分离人源DNA片段中的应用

Identification and isolation of the human anonymous DNAs by Inter Alu PCR

目的：探讨ＩｎｔｅｒＡｌｕＰＣＲ技术在筛选鉴定含人ＤＮＡ小鼠转染细胞和分离人源ＤＮＡ片段中的应用。方法：设计人特异性Ａｌｕ引物，通过ＩｎｔｅｒＡｌｕＰＣＲ扩增对转染人基因组ＤＮＡ后回复突变的小鼠细胞进行鉴定筛选；分离插入的人源ＤＮＡ片段；经原位ＰＣＲ鉴定其人源性。结果：应用该技术从８株回复突变株中筛选出６株含特异性人ＤＮＡ；从其中１２号小鼠细胞基因组ＤＮＡ中分离出３条人源ＤＮＡ片段；ＰＣＲ原位杂交确证所分离的ＤＮＡ片段具有人的特异性。结论：ＩｎｔｅｒＡｌｕＰＣＲ技术是应用于筛选鉴定含人ＤＮＡ杂种细胞、分离其中人源插入片段的简便、有效的方法。

Objective: To identify the mouse tumor cell clones transfected with normal human genomic DNAs and to isolate the inserted human anonymous sequences by Inter Alu PCR. Methods: Human DNA fragments were amplified by Inter Alu PCR technique from normal human genomic DNAtransfected mouse tumor cell clones, of which the metastatic phenotype had been suppressed. The human origin of the amplified DNA was further confirmed by PCR in situ hybridization. Results: From eight revertants exhibiting normal phenotype, six clones containing human DNA were identified by Inter Alu PCR ; a continuum of human genomic DNA fragments was obtained from number 12 mouse tumor cell, of which the metastatic phenotype was suppressed. Conclusion: The results indicated that Inter Alu PCR is a specific technique to amplify human gene sequences from complex heterologous DNAs by using human Alu primers.