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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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CRISPR and crRNAs02:53

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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.
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Genomics02:02

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Genomics is the science of genomes: it is the study of all the genetic material of an organism. In humans, the genome consists of information carried in 23 pairs of chromosomes in the nucleus, as well as mitochondrial DNA. In genomics, both coding and non-coding DNA is sequenced and analyzed. Genomics allows a better understanding of all living things, their evolution, and their diversity. It has a myriad of uses: for example, to build phylogenetic trees, to improve productivity and...
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While every living organism has a genome of some kind (be it RNA, or DNA), there is considerable variation in the sizes of these blueprints. One major factor that impacts genome size is whether the organism is prokaryotic or eukaryotic. In prokaryotes, the genome contains little to no non-coding sequence, such that genes are tightly clustered in groups or operons sequentially along the chromosome. Conversely, the genes in eukaryotes are punctuated by long stretches of non-coding sequence.
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Genome Engineering of Primary Human B Cells Using CRISPR/Cas9
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Optimizing a CRISPR-Cpf1-based genome engineering system for Corynebacterium glutamicum.

Jiao Zhang1, Fayu Yang1, Yunpeng Yang1,2

  • 1Key Laboratory of Molecular Medicine and Biotherapy, School of Life Sciences, Beijing Institute of Technology, No. 5 South Zhongguancun Street, Beijing, 100081, China.

Microbial Cell Factories
|March 27, 2019
PubMed
Summary
This summary is machine-generated.

Optimizing the CRISPR-Cpf1 system in Corynebacterium glutamicum enhances genome editing efficiency. This study identified key crRNA parameters, including a 5'-NYTV-3' PAM and 21 bp spacer, to improve targeting for industrial applications.

Keywords:
CRISPR-Cpf1Corynebacterium glutamicumIsobutyrateLinear templatePAMcrRNA

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Area of Science:

  • Microbiology
  • Molecular Biology
  • Biotechnology

Background:

  • Corynebacterium glutamicum is a key industrial microorganism for chemical production.
  • CRISPR-Cpf1 systems offer programmable genome editing but face efficiency challenges in C. glutamicum.
  • Low editing efficiency (<15%) in C. glutamicum is linked to poor crRNA targeting.

Purpose of the Study:

  • To develop a screening strategy for evaluating crRNA targeting efficiency in C. glutamicum.
  • To optimize the CRISPR-Cpf1 system for enhanced genome editing in C. glutamicum.
  • To improve the production of valuable chemicals using engineered C. glutamicum.

Main Methods:

  • Systematic screening of crRNA targeting efficiency in C. glutamicum.
  • Quantitative analysis of CRISPR-Cpf1 system parameters (PAM, spacer length, repair template).
  • Evaluation of linear DNA for double-strand break repair.

Main Results:

  • Identified an optimal crRNA with a 5 extbackslash'NYTV-3 extbackslash' PAM and 21 bp spacer for high targeting efficiency.
  • Demonstrated that linear DNA can effectively repair double-strand breaks in C. glutamicum.
  • Achieved higher genome editing efficiency compared to previous methods.

Conclusions:

  • Optimized PAM and crRNA parameters significantly enhance the CRISPR-Cpf1 system in C. glutamicum.
  • The findings provide insights into FnCpf1 endonuclease function and Cpf1-based genome editing.
  • The improved system holds potential for increasing the production of bulk chemicals like isobutyrate.