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Related Concept Videos

CRISPR/Cas9 Genome Editing01:28

CRISPR/Cas9 Genome Editing

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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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CRISPR01:59

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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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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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RNA Editing02:23

RNA Editing

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RNA editing is a post-transcriptional modification where a precursor mRNA (pre-mRNA) nucleotide sequence is changed by base insertion, deletion, or modification. The extent of RNA editing varies from a few hundred bases, in mitochondrial DNA of trypanosomes, to a just single base, in nuclear genes of mammals. Even a single base change in the pre-mRNA can convert a codon for one amino acid into the codon for another amino acid or a stop codon. This type of re-coding can significantly affect the...
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Genomics02:02

Genomics

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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 Editing in Mammalian Cell Lines using CRISPR-Cas
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CRIS.py: A Versatile and High-throughput Analysis Program for CRISPR-based Genome Editing.

Jon P Connelly1,2, Shondra M Pruett-Miller3,4

  • 1St. Jude Children's Research Hospital, Department of Cell & Molecular Biology, Memphis, 38105, USA. patrick.connelly@stjude.org.

Scientific Reports
|March 14, 2019
PubMed
Summary
This summary is machine-generated.

CRISPR-Cas9 gene editing efficiency can now be easily quantified using CRIS.py, a new Python tool. This program analyzes next-generation sequencing data to identify gene knock-outs and knock-ins across many samples simultaneously.

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Area of Science:

  • Genomics
  • Molecular Biology
  • Bioinformatics

Background:

  • CRISPR-Cas9 enables precise genomic modifications but quantifying editing efficiency and identifying edited clones is challenging.
  • Targeted next-generation sequencing (NGS) offers high-throughput analysis but generates large datasets difficult to interpret.
  • Existing NGS analysis tools for CRISPR editing have limitations in scalability and flexibility.

Purpose of the Study:

  • To develop a versatile Python-based program, CRIS.py, for analyzing CRISPR-Cas9 editing outcomes.
  • To provide a streamlined method for quantifying gene editing rates and identifying modified clones from NGS data.
  • To offer an efficient solution for analyzing large-scale CRISPR editing experiments.

Main Methods:

  • Development of a Python program, CRIS.py, for automated analysis of NGS data.
  • Implementation of algorithms to concurrently detect knock-out and multiple user-specified knock-in modifications.
  • Design of an output format for easy searching and identification of correctly edited samples.

Main Results:

  • CRIS.py can analyze thousands of samples simultaneously, significantly improving throughput.
  • The program accurately identifies both knock-out and multiple knock-in events, including base editor-induced modifications.
  • CRIS.py provides an easily searchable output, simplifying the identification of desired clones.

Conclusions:

  • CRIS.py offers a significant advancement in analyzing CRISPR-Cas9 editing efficiency from NGS data.
  • The tool enhances the ability to optimize editing reagents and identify correctly modified clones.
  • CRIS.py provides a scalable, versatile, and user-friendly solution for CRISPR editing analysis.