郜刚分子生物学-03-染色体-2包装和半保留复制.ppt

Transmission electron micrograph of human a HeLa cell, illustrating the replication bubble that characterizes DNA replication within a single replicon. A Picture Paints a Thousand Words... or more? Because a figure is as informative as 1000 words…. DNA Replication Mode （3）Semiconservative replication（半保留复制） The expected results of two generations of semiconservative replication in the Meselson-Stahl experiment.. Meselson－Stahl实验的改进之一 对15N-DNA、14N-DNA杂交分子经加热变性，对于变性前后的DNA分别进行CsCl密度梯度离心。 结果变性前的杂交分子为一条中间密度带，变性后则分为两条区带，即重密度带（15N-DNA）和低密度带（14N-DNA）。 第0代 只有15N，第一代出现杂合分子，两代后出现等量的14N和14N-15N杂合分子。纯粹15N的分子没有的了，说明老链保留到了新链中了。 Semi-conservative replication of chromosomes can also be visualized through an examination of chromosomes that are allowed two rounds of replication in a medium containing bromodeoxy-uridine. These replicated chromosomes are then stained with a fluorescent dye and Giemsa stain to produce what are described as harlequin chromosomes DNA半保留复制的生物学意义 DNA的半保留复制表明DNA在代谢上的稳定性，保证亲代的遗传信息稳定地传递给后代。 Multiple Origins in Eukaryotes 2.3.3复制的几种主要方式page40 “复制方式”不同于“复制机制” 线性DNA的复制： 典型的复制叉式的复制 环状DNA的复制： θ复制 滚环复制 D环复制 2. 环状DNA双链的复制 John Cairns’ experiment In 1963, John Cairns: Grew E. coli in 3H thymidine Waited till cells were in the middle of replication Lysed the cells very very gently Spread the lysate on an EM grid Exposed the grid to X-ray film for TWO months Replication of the E. coli chromosome what Cairns’ experiment showed:θ E. coli has a circular chromosome E. coli has a single origin of replication In E. coli, replication and unwinding are simultaneous What Cairns did not show: ? Is replication unidirectional or bidirectional? ? Is the origin of replication unique? ? Where is the origin of replication located? θ复制的特点 多数环状DNA采用的复制方式 复制叉式复制，两个复制叉，双向等速或者不等速复制 有先导链和后随链之分 滚环复制 滚环型复制： 单向复制的特殊方式 T. Brown et al., 如：ΦΧ174的双链环状DNA复制型（RF） （1）模板链和新合成的链分开; （2）不需RNA引物，新生链在正链3‘－OH上共价延伸延伸 （3）只有一个复制叉;复制是单向的 (4)滚环复制也有