基因组改造解说.ppt

构建人造染色体： 需要与着丝粒功能有关的alphoid DNA 以及端粒序列 RFLP操作十分简单，即选择一个可以与目标区域杂交的探针进行Southern blotting分析。 在高分辨遗传和物理图谱被建立之前，利用RFLP已经使得很多基因被标注到了它们在染色体上的位置上: Huntington disease (4号染色体) 囊性纤维化（cystic fibrosis, 7号染色体） cystic fibrosis 镰形血球贫血病（11号染色体） sickle-cell anaemia 视网膜母细胞瘤（13号染色体） retinoblastoma 阿尔兹海默氏症（21号染色体） Alzheimer disease 使用RFLP标记，在1987年产生了第一张人类基因组遗传图谱。 RFLP方法的多态性程度和分辨能力十分有限，一个限制性位点只能是存在或缺少两种形式。 小卫星DNA和微卫星DNA的使用使得构建更为详尽的遗传图谱成为可能. minisatellites and microsatellites Using RFLPs as markers, a genetic map of the human genome was available by 1987. However, this approach was limited in terms of the degree of polymorphism (a restriction site can only be present or absent) and in the level of resolution. 小卫星由短序列（10-100个碱基）串联重复组成，小卫星区域的重复单位数目是可以变化的。 又叫做可变数目串联重复（VNTRs, variable number tandem repeats）. minisatellites are made up of tandem repeats of short (10-100 base-pair) sequences. The number of elements in a minisatellite region can vary, and thus these are also known as variable number tandem repeats, VNTRs. VNTR的用途：作图研究、遗传指纹分析 VNTR在染色体上分布不均匀，倾向于分布在染色体末端； VNTRs can be used in mapping studies, and also formed the basis of genetic fingerprinting. One drawback with VNTRs is that they are not evenly distributed in the genome, tending to be located at the ends of chromosomes. 微卫星是一些重复单位更短的重复序列，在作图研究中CA重复已被用作标准微卫星DNA。 STR，short tandem repeat. CA重复区域可以用PCR进行扩增，引物序列位于重复序列两翼。 Microsatellites are much shorter repeats, and the two base-pair CA repeat has been used as the standard microsatellite in mapping studies. Regions of CA repeats can be amplified using PCR, with primers that flank the repeated elements. 由于引物来自单一序列区，所以通过这种办法扩增的微卫星属于STS的一种类型。 可以用于将遗传图谱和物理图谱联系起来. As the primers are derived from unique-sequence regions, this essentially means that microsatellites amplified in this way are a type of STS, and can therefore be used to link genetic and physical maps. 1995年发表的物理图谱含有15000个STS标记。 1998年增加到30000个物理标记。 A physical map with over 15000 STS markers was publishe