Difference between revisions of "Part:BBa K4088898"

Latest revision as of 02:11, 22 October 2021

dCas13a

Cas13a is a classic RNA-targeting nuclease that is widely used in a variety of diagnostics methods. It is one enzyme of Cas13 family which contains at least 4 subtypes, including Cas13a, used as our Parts BioBrick. We took LwaCas13a, identified from Leptotrichia wadei (Lwa).

dCas13a is a catalytically dead lwaCas13a enzyme (formerly C2c2) with R407A and R1046A mutations in HEPN-domain. dCas13a interacts with gRNA about 65 nucleotides in length. dCas13a-gRNA complex detects complementary RNA in the sample. dCas13a is very specific variant of RNA detection, it can be customized to recognize single nucleotides on the target RNA.

LwaCas13a has been reported to mediate more robust RNA-targeting activity than other Cas13 systems like LshCas13, but it requires a stabilizer fusion, for example, msfGFP for efficient interference activity ^[1]. As a stabilizer fusion we use fragments of beta-lactamase thus improving targeting activity.
For more information, see our part BBa_K4088891.

Sequence and Features

Assembly Compatibility:

10
INCOMPATIBLE WITH RFC[10]
Illegal EcoRI site found at 622
Illegal PstI site found at 601
12
INCOMPATIBLE WITH RFC[12]
Illegal EcoRI site found at 622
Illegal PstI site found at 601
21
INCOMPATIBLE WITH RFC[21]
Illegal EcoRI site found at 622
Illegal BglII site found at 924
Illegal BglII site found at 1689
Illegal BglII site found at 1956
Illegal BglII site found at 2166
23
INCOMPATIBLE WITH RFC[23]
Illegal EcoRI site found at 622
Illegal PstI site found at 601
25
INCOMPATIBLE WITH RFC[25]
Illegal EcoRI site found at 622
Illegal PstI site found at 601
1000
COMPATIBLE WITH RFC[1000]

↑ Mahas, A., Aman, R. & Mahfouz, M. CRISPR-Cas13d mediates robust RNA virus interference in plants. Genome Biol 20, 263 (2019). https://doi.org/10.1186/s13059-019-1881-2

[1] Mahas, A., Aman, R. & Mahfouz, M. CRISPR-Cas13d mediates robust RNA virus interference in plants. Genome Biol 20, 263 (2019). https://doi.org/10.1186/s13059-019-1881-2

[1]

@@ Line 3: / Line 3: @@
 <partinfo>BBa_K4088898 short</partinfo>
-Cas13a is a classic RNA-targeting nuclease that is widely used in a variety of diagnostics methods. It is a very specific variant of RNA detection, it can be customized to recognize single nucleotides on the target RNA. dCas13a is a catalytically dead LwaCas13a enzyme.<br/>
+Cas13a is a classic RNA-targeting nuclease that is widely used in a variety of diagnostics methods. It is one enzyme of Cas13 family which contains at least 4 subtypes, including Cas13a, used as our Parts BioBrick. We took LwaCas13a, identified from Leptotrichia wadei (Lwa).<br/>
+dCas13a is a catalytically dead lwaCas13a enzyme (formerly C2c2) with R407A and R1046A mutations in HEPN-domain. dCas13a interacts with gRNA about 65 nucleotides in length. dCas13a-gRNA complex detects complementary RNA in the sample. dCas13a is very specific variant of RNA detection, it can be customized to recognize single nucleotides on the target RNA.<br/>
+LwaCas13a has been reported to mediate more robust RNA-targeting activity than other Cas13 systems like LshCas13, but it requires a stabilizer fusion, for example, msfGFP for efficient interference activity <ref> Mahas, A., Aman, R. & Mahfouz, M. CRISPR-Cas13d mediates robust RNA virus interference in plants. Genome Biol 20, 263 (2019). https://doi.org/10.1186/s13059-019-1881-2</ref>. As a stabilizer fusion we use fragments of beta-lactamase thus improving targeting activity.<br/>
 For more information, see our part <partinfo>BBa_K4088891</partinfo>.
 <br/><br/>