CRISPR-Cas12a is a genome editing tool similar to CRISPR-Cas9. Both systems use guide RNA, which are designed by the researcher, to target the protein complex to DNA and both produce targeted, site-specific double stranded DNA (dsDNA) breaks. CRISPR-Cas12a is more precise and less tolerant of mismatches between the guide RNA and target sequence than CRISPR-Cas9 .
In addition to this activity, Jennifer Doudna’s lab at University of California, Berkeley and Jin Wang’s lab at Chinese Academy of Sciences, Shanghai reported that CRISPR-Cas12a indiscriminately degrades single-stranded DNA (ssDNA) in trans to ssDNA not complimentary to the guide RNA . Wang’s lab also described guide RNA directed ssDNA cleavage for CRISPR-Cas12a.
Since ssDNA degradation in trans is activated by CRISPR-Cas12a recognition of target DNA, Doudna’s group developed the DNA Endonuclease Targeted CRISPR Trans Reporter (DETECTR) system to detect specific DNA sequences like dsDNA viruses . The system includes a ssDNA fluorophore quencher that fluoresces upon ssDNA degradation to report detection of the HPV DNA sequence by CRISPR-Cas12a. DETECTR detected human papillomavirus (HPV) and distinguished between two types of HPV in DNA extracted from human anal swabs.
Wang’s group developed a similar system called HOur Low-cost Multipurpose highly Efficient System (HOMES) to detect DNA sequences using CRISPR-Cas12a with a quenched ssDNA reporter . In extracted DNA from human cell lines or PCR amplified saliva samples HOMES distinguished sequence variations between samples at single nucleotide polymorphism (SNP) sites that are related to human health and personal characteristics. HOMES also detected viruses and distinguished between different viral strains. To detect RNA viruses, RNA samples need to be first reverse transcribed into cDNA.
The CRISPR-Cas12a non-specific ssDNA degradation is similar to the non-specific cleavage of RNA observed by CRISPR-Cas13a. Similarly, non-specific cleavage of RNA is used as a readout for when CRISPR-Cas13a detects viral RNA sequences in the SHERLOCK system.
The gene editing technology company Inscripta has developed a CRISPR enzyme similar to Cas12a called MAD7.
Characterized CRISPR-Cas12 activity, developed DETECTR
Characterized CRISPR-Cas12 activity developed HOMES
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- Cluster: Synthetic biologyA cluster of topics related to synthetic biology.
- CRISPRClustered regularly interspaced short palindromic repeats (CRISPR) is a prokaryotic adaptive immune response that provides immunity against foreign nucleic acids, such as viral DNA and bacterial plasmids, through the use of crRNAs (CRISPR RNAs) and associated Cas genes.
- CRISPR/Cas ToolsClustered regularly interspaced short palindromic repeats (CRISPR) and CRISPR associated (Cas) proteins perform adaptive immune functions in prokaryotic organisms defending against foreign nucleic acids such as viruses. CRISPR/Cas tools have been adapted for use in genome editing and other DNA and RNA targeting applications.
- Synthetic biologyInterdisciplinary branch of biology and engineering, applying multiple disciplines to build artificial biological systems for research, engineering, and medical applications.
- Gene editing
- DNAMolecule that encodes the genetic instructions used in the development and functioning of all known living organisms and many viruses.
- CRISPR-Cas13A nucleic acid editing technology that targets RNA, analogous to the CRISPR-Cas9 system
- CRISPR-Cas12dCRISPR-Cas12d belongs to the Type V CRISPR systems that confer bacterial immunity against viruses and contains the Class II effector protein Cas12d.
- InscriptaA gene editing technology company founded in 2015 and based in Boulder, CO, USA.
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