Goldammer T, Weikard R, Brunner R M, Schwerin M
Forschungsbereich Molekularbiologie, Forschungsinstitut für die Biologie landwirtschaftlicher Nutztiere, D 18196 Dummerstorf, Germany.
Mamm Genome. 1996 Apr;7(4):291-6. doi: 10.1007/s003359900085.
A rapid procedure for the defined isolation and characterization of single bovine chromosome fragment specific probes is described. This has been developed as a technical prerequisite for the directed generation of bovine DNA sequences. The specific regions 1q13-24, 5q21-24, 6q31-32, 7q21-22, 12q24-ter, and 20q12-ter of bovine GTG-banded metaphase chromosomes were microdissected and amplified by PCR with a degenerate oligonucleotide primer and subsequently cloned into pBluescript II SK. The DNA probes generated were characterized by gel electrophoresis, dot blot analysis and rehybridization in situ to GTG-banded metaphase spreads. The position and size of the hybridization sites on the chromosomes correspond exactly to the dissected chromosome areas and indicate the complexity and specificity of the microdissected and amplified chromosome material.
本文描述了一种用于明确分离和鉴定单个牛染色体片段特异性探针的快速方法。这一方法是作为定向生成牛DNA序列的技术前提而开发的。对牛GTG带型中期染色体的特定区域1q13 - 24、5q21 - 24、6q31 - 32、7q21 - 22、12q24 - ter和20q12 - ter进行显微切割，并用简并寡核苷酸引物通过PCR扩增，随后克隆到pBluescript II SK中。通过凝胶电泳、斑点印迹分析以及与GTG带型中期染色体铺展进行原位再杂交，对所产生的DNA探针进行了鉴定。染色体上杂交位点的位置和大小与显微切割的染色体区域完全对应，表明显微切割和扩增的染色体材料具有复杂性和特异性。
