热启动PCR快速制备地高辛标记探针

介绍了一种在热启动PCR中,以Dig-11-dUTP部分代替dTTP,从少量基因组DNA中快速制备大量的地高辛标记的探针的方法,此探针灵敏度达0.03pg,并只和相关的DNA特异杂交.     

Quox-1基因同源序列在小鼠精子发生中的表达

Ｑｕｏｘ－１基因是从鹌鹑中分离得到的Ａｎｔｐ类型的一个同源异形盒基因。以＾３２Ｐ标记的Ｑｕｏｘ－１基因的ｃ３片段为探针,采用分子杂交技术确定了小鼠基因组中存在Ｑｕｏｘ－１基因同源序列。以抗ＱＵＯＸ－１蛋白的特异性抗体对幼年小鼠睾丸、成年小鼠睾丸及附睾的蛋白质样品和组织切片,分别进行了Ｗｅｓｔｅｒｎ ｂｌｏｔ分析和免疫组织化学反应。结果证明,性成熟小鼠在精子发生过程中的精子形成阶段有类ＱＵＯＸ－１蛋     

Phylogenetic relationships among six species of Epistylis inferred from 18S-ITS1 sequences

Phylogenetic relationships among six species of Epistylis (i. e. E. plicatilis, E. urceolata, E. chrysemydis, E. hentscheli, E. wenrichi, and E. galea) were investigated using sequences of the first internal transcribed spacer region (ITS-1) of ribosomal DNA (rDNA). Amplified rDNA fragment sequences consisted of 215 or 217 bases of the flanking 18S and 5.8S regions, and the entire ITS-1 region (from 145 to 155 bases). There were more than 33 variable bases between E. galea and the other five species in both the 18S region and the ITS-1 region. The affiliation of them was assessed using Neighbor-joining (NJ), maximum parsimony (MP) and maximum likelihood (ML) analyses. In all the NJ, MP and ML analyses E. galea, whose macronucleic position and shape are distinctly different from those of the other five species, was probably diverged from the ancestor of Epistylis earlier than the other five species. The topology in which E. plicatilis and E. hentscheli formed a strongly supported sister clade to E. urceol     

In situ amplification of DNA fragments specific for human Y chromosome in cellular nuclei by PCR

Using single primer pairs Y3 and Y4, in siru polymerase chain reaction (in situ PCR) was successfully performed on the specimen slides of peripheral leukocytes. By both of the direct digpxiginin-11-dUTP incorporation into PCR products with in situ PCR (direct in situ PCR) and in situ PCR followed by detection of in situ hybridization (indirect in siru PCR), DNA fragments specific for human Y chromosome were obviously amplified in cellular nuclei of specimens on the slides. The results were verified by Southern analysis. The methodology of in situ PCR and its application were discussed.     

人Y染色体特异DNA片段的原位PCR扩增

报道了以人类外周血白细胞为材料,利用人类Y染色体特异片段一对扩增引物Y3和 Y₄,在玻片细胞样品上进行原位 PCR获得的结果.用Dig-11-dUTP于原位PCR过程中直接掺入法(直接原位PCR)和细胞原位PCR后再作原位杂交的方法(间接原位PCR)都证明:玻片上细胞样品内的Y染色体特异DNA片段有明显的原位扩增.并对原位PCR的专一性与可重复性,细胞内PCR扩增产物的存留等方法学问题,以及原位PCR技术的价值和应用前景进行了讨论.