Week of 4/29

 Introduction

This week was spent on researching and beginning the design for my own project, to be started in the upcoming fall semester. I have been asked to turn my focus towards CRISPR. In recent experiments, D. xinjiangensis failed to withstand ultraviolent radiation at a concentration of 30000µJ/cm2 . My idea is to utilize CRISPR to edit the genes of D.xinjiangensis to have a more efficient DNA repair mechanism.


Body

CRISPR DNA Repair Notes

- homologous recombination: nucleotide sequences are exchanged between two identical DNA molecules (this is a DNA method)

- mutagenesis: process by which an organism's DNA changes, resulting in a mutation

-DDR- DNA damage response

- PprI-DdrO:

a. DdrO represses transcription of DDR genes before stresses occur

b. PprI is activated after sensing of DNA damage signals

c. PprI cleaves DdrO through an unclear mechanism

d. DDR genes unregulated until damage is repared

- Proteins involved in Deinoccoccus DNA damage response: PprI,PprA,DrRA

-CRISPR/CAS system has 2 modules: recognition and cleavage

-recognition module: recognition module sequence is transcribed into a single-guide RNA (sgRNA) capable of targeting the genome

-cleavage module: CAS endonuclease forms a ribonucleoprotein complex with sgRNA (recognizes a short, conserved sequence known as protospacer adjacent motif or PAM)

-CAS endonuclease introduces a double-strand break (DSB) upon recognition of sequence complementary to sgRNA sequences adjacent to PAM sequence, can be directly repaired by endogenous DNA repair pathways

Procedure

The following flowchart describes the first, very rough draft of the project's procedure:

Resources:

Precise CRISPR/Cpf1 genome editing system in the Deinococcus radiodurans with superior DNA repair mechanisms - ScienceDirect


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