Lecture 13: CRISPR
Segment 1: Introduction to CRISPR
Segment 2: Review of Homologous Recombination
Segment 3: 真核生物中的同源重组Homologous Recombination in Eukaryotes
Segment 4: Cell Cycle Terms
概览一些生物学中重要的术语和概念
Segment 5: Cell Cycle
Segment 6: 减数分裂重组Meiotic Recombination, Part 1
宏观上将减数分裂导致的DNA交换
Segment 7: 减数分裂重组Meiotic Recombination, Part 2
从微观上讲
Segment 8: Genome Editing and CRISPR, Part 1
CRISPR：Clustered Regularly Interspaced Short Palindromic Repeats
class I 需要多个蛋白；而class II只需要1个蛋白，比如下图中的cas9 蛋白，一个就能定位靶基因。而且cas gene恰好跟repeat region相邻，只要找到了repeats,就能找到设计的蛋白
Segment 9: Genome Editing and CRISPR, Part 2
为什么不会定向到细菌自己的基因呢？有两个解答，现仅以一个为例：
如下图所示，细菌转录本包含一段spacer（黄色）和一段repeat（灰色），左边有tracrRNA以反式激活CRISPR RNA，若tracrRNA不表达，则系统不工作。Cas9 蛋白实现剪切，Cas1和Cas2蛋白从噬菌体中获得新的序列，PAM是2-3个核苷酸长度的motif。因为细菌中没有PAM，所以不会被CRISPR/Cas9系统所剪切
Segment 10: Genome Editing and CRISPR, Part 3
下图定位于target genome中，PAM是NGG，此图中的N为C，黄色的序列是上图中的protospacer，又称作guide RNA，而灰色的是repeat序列，又称作linker。蓝色的是tracrRNA。所以可以通过改变guide RNA 来定向识别想要编辑的基因。箭头所指向的是Nuclease Sites核酸酶位点。
You wish to use Cas9 to activate expression of a gene of interest. How should you modify your CRISPR gene?
- mutate the gene encoding Cas9 to eliminate the nuclease activity核酸酶活性，这样就不会切割目的基因了。
- Express Cas9 as a fusion protein融合蛋白 with a transcriptional activator转录激活因子。这样可以将转录激活因子带到想要表达的基因处，以激活该基因。
Additional CRISPR Video Resources from Our Colleagues
What is CRISPR? (2014)
Feng Zhang Explains CRISPR
Feng Zhang Lecture
CRISPR Applications to Human Health: SHERLOCK
一些文章
Dynamics and Impact of Homologous Recombination on the Evolution of Legionella pneumophila
CRISPR/Cas9 Targeting Events Cause Complex Deletions and Insertions at 17 sites in the Mouse Genome
CRISPR/Cas9-mediated Mutation of Tyrosinase (Tyr) 3′ UTR Induce Graying in Rabbit
CRISPR/Cas9-APEX-Mediated Proximity Labeling Enables Discovery of Proteins Associated with a Predefined Genomic Locus in Living Cells
