摘要
反向斑点杂交是将寡核苷酸探针固定在膜上，用标记的靶序列与固定在膜上的探针进行杂交．与正向杂交相比，它通过一次杂交即可确定多种基因型，是一种快速筛查ＤＮＡ点突变的诊断方法我们选择β－地中海贫血基因－２８（Ａ→Ｇ）、ＣＤ１７（Ａ→Ｔ）、ＣＤ４１～４２（－ＴＴＣＴ）三种点突变为模型采用ＰＣＲ扩增产生串联多拷贝序列探针并将其克隆化．将克隆化探针固定于尼龙膜上，与同位素标记的β－珠蛋白基因ＰＣＲ片段进行杂交，检测β－地贫患者的基因型．
Reverse dot blot is featured by hybridization of labeled target DNA with immobilized probes. In conventional dot blot, each probe requires a separate hybridization, while in reverse dot blot all known allelic variants at several loci in a sample can be screened simultaneously. PCR was used to generate tandem polymers of ASO sequences for three β-thalassemia mutations [-28 (A→ G), CD17 (A→T) and CD41～42 (-TTCT) ], and then inserted the amplified probe polymers into plasmid. The final probes, which were amplified by PCR, were applied to the nylon membrane and hybridized with radioactive labeled HBB PCR products to detect the genotype of sample DNA.
出处
《生物化学与生物物理进展》
SCIE
CAS
CSCD
北大核心
1996年第3期273-278,共6页
Progress In Biochemistry and Biophysics
