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Related Concept Videos

CRISPR01:59

CRISPR

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Genome editing technologies allow scientists to modify an organism’s DNA via the addition, removal, or rearrangement of genetic material at specific genomic locations. These types of techniques could potentially be used to cure genetic disorders such as hemophilia and sickle cell anemia. One popular and widely used DNA-editing research tool that could lead to safe and effective cures for genetic disorders is the CRISPR-Cas9 system. CRISPR-Cas9 stands for Clustered Regularly Interspaced...
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The CRISPR-Cas system serves as a bacterial defense mechanism against invading genetic elements such as viruses and plasmids, forming the foundation for its adaptation as a powerful genome-editing tool. Originally discovered in prokaryotes, this system has been repurposed to revolutionize genetic engineering across a wide range of organisms, including plants, animals, and humans. The core component, Cas9, is an endonuclease derived from Streptococcus pyogenes, capable of introducing...
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Combinatorial Gene Control02:33

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Combinatorial gene control is the synergistic action of several transcriptional factors to regulate the expression of a single gene. The absence of one or more of these factors may lead to a significant difference in the level of gene expression or repression.
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CRISPR and crRNAs02:53

CRISPR and crRNAs

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Bacteria and archaea are susceptible to viral infections just like eukaryotes; therefore, they have developed a unique adaptive immune system to protect themselves. Clustered regularly interspaced short palindromic repeats and CRISPR-associated proteins (CRISPR-Cas) are present in more than 45% of known bacteria and 90% of known archaea.
The CRISPR-Cas system stores a copy of foreign DNA in the host genome and uses it to identify the foreign DNA upon reinfection. CRISPR-Cas has three different...
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Regulation of Expression at Multiple Steps01:23

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The gene expression in cells is regulated at different stages: (i) transcription, (ii) RNA processing, (iii) RNA localization, and (iv) translation. Transcriptional regulation is mediated by regulatory proteins such as transcription factors, activators, or repressors—these control gene expression by initiating or inhibiting the transcription of genes. Once a precursor or pre-mRNA is produced, it undergoes post-transcriptional modification, including 5' capping, splicing, and the...
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Gene expression can be regulated at almost every step from gene to protein. Transcription is the step that is most commonly regulated. This involves the binding of proteins to short regulatory sequences on the DNA. This association can either promote or inhibit the transcription of a gene associated with the respective sequence.
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Gene Digital Circuits Based on CRISPR-Cas Systems and Anti-CRISPR Proteins
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Multiplex gene regulation by CRISPR-ddCpf1.

Xiaochun Zhang1,2,3, Jingman Wang4,5, Qiuxiang Cheng6

  • 1Key Laboratory of Synthetic Biology, Institute of Plant Physiology and Ecology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai, China.

Cell Discovery
|June 14, 2017
PubMed
Summary
This summary is machine-generated.

The CRISPR-ddCpf1 system enables multiplex gene regulation in E. coli by processing multiple CRISPR RNAs (crRNAs) efficiently. This novel approach simplifies targeting multiple genes for research and potential clinical applications.

Keywords:
CRISPRCRISPRiCpf1DNase-dead Cpf1 (ddCpf1)multiplex gene regulation

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Area of Science:

  • Molecular Biology
  • Microbial Genetics

Background:

  • The Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)/dCas9 system is valuable for transcriptional and epigenetic studies.
  • Multiplex gene targeting traditionally requires independent expression of numerous single-guide RNAs (sgRNAs), posing practical challenges.

Purpose of the Study:

  • To develop a more convenient method for multiplex gene regulation using a DNase-dead Cpf1 (ddCpf1) mutant.
  • To investigate the efficacy and specificity of ddCpf1-mediated gene repression in *Escherichia coli*.

Main Methods:

  • Utilized a DNase-dead Cpf1 (ddCpf1) mutant for multiplex gene regulation in *E. coli*.
  • Employed whole-transcriptome RNA-sequencing to assess the specificity of ddCpf1-mediated repression.
  • Demonstrated *in vivo* processing of precursor CRISPR arrays into mature CRISPR RNAs (crRNAs) by ddCpf1.

Main Results:

  • ddCpf1 mediated gene repression in *E. coli*, with enhanced efficacy when crRNAs targeted the template strand, differing from dCas9.
  • Both DNA strands were effectively repressed by the ddCpf1/crRNA complex when targeting promoter regions.
  • ddCpf1's residual RNase activity facilitated the generation of multiple mature crRNAs from a precursor array, enabling multiplex gene regulation.
  • The system was successfully applied to rapidly screen a library of two-component systems in *E. coli*.

Conclusions:

  • The CRISPR-ddCpf1 system offers a simplified and effective strategy for multiplex gene regulation.
  • This system demonstrates high specificity and facilitates rapid screening of gene targets.
  • The CRISPR-ddCpf1 system holds potential for broad applications in biological research and clinical studies.