Difference between revisions of "Part:BBa K4367014"

Latest revision as of 13:01, 12 October 2022

ABI-cTEV
ABI-cTEV is a part of the ABI/PYL-splitTEV system, which is used as a control for other experiments. Essentially, the complementation of split TEV can be induced with the addition of abscisic acid to the system.

Description

The design of ABI-cTEV is essentially identical to what was used in this reference [1], but with the addition of a 6xHis-tag and Part-3-Overhangs for Modular Cloning. PYL and ABI can dimerize if abscisic acid is present [1]. When PYL-nTEV dimerises with ABI-cTEV, the split TEVp halves can complement each other and regain enzymatic function. More information can be found at BBa_K4367010 (TEVp).

Usage

It was planned to use the ABI/PYL-splitTEV system as a control for the dCas9-splitTEV system, as stepping stones for troubleshooting which parts could be dysfunctional. For example, if the ABI/PYL system works but not the dCas9-splitTEV system, it is shown that the testing conditions can allow for complementation of split TEV, and the problem is perhaps because of the dCas9 or gRNA.

Future design considerations

If for some reason the ABI/PYL system would not be desirable, there is an alternative: The proteins FKBP and FRB, which uses the chemical rapamycin to induce their dimerisation [1].

Sequence and Features

Assembly Compatibility:

10
COMPATIBLE WITH RFC[10]
12
COMPATIBLE WITH RFC[12]
21
INCOMPATIBLE WITH RFC[21]
Illegal BamHI site found at 952
Illegal BamHI site found at 1034
Illegal BamHI site found at 1363
23
COMPATIBLE WITH RFC[23]
25
COMPATIBLE WITH RFC[25]
1000
INCOMPATIBLE WITH RFC[1000]
Illegal BsaI site found at 14
Illegal BsaI.rc site found at 1370
Illegal SapI.rc site found at 1270

References

[1] Tina Fink, Jan Lonzarić, Arne Praznik, Tjaša Plaper, Estera Merljak, Katja Leben, Nina Jerala, Tina Lebar, Žiga Strmšek, Fabio Lapenta, Mojca Benčina & Roman Jerala (2019). Design of fast proteolysis-based signaling and logic circuits in mammalian cells. Available at: https://www.nature.com/articles/s41589-018-0181-6

@@ Line 6: / Line 6: @@
 <h2>Description</h2>
-The design of ABI-cTEV is essentially identical to what was used in this reference [1], but with the addition of a 6xHis-tag and Part-3-Overhangs for Modular Cloning. PYL and ABI can dimerize if abscisic acid is present [1]. When PYL-nTEV dimerises with ABI-cTEV, the split TEV can complement each other and regain enzymatic function. More information can be found at BBa_K4367010 (TEVp).
+The design of ABI-cTEV is essentially identical to what was used in this reference [1], but with the addition of a 6xHis-tag and Part-3-Overhangs for Modular Cloning. PYL and ABI can dimerize if abscisic acid is present [1]. When PYL-nTEV dimerises with ABI-cTEV, the split TEVp halves can complement each other and regain enzymatic function. More information can be found at BBa_K4367010 (TEVp).
 <h2>Usage</h2>
-It was planned to use the ABI/PYL-splitTEV system as a control for the dCas9-splitTEV system, and to use as stepping stone for troubleshooting what parts could be dysfunctional. For example, if the ABI/PYL system works but not the dCas9-splitTEV system, it's shown that the testing conditions can allow for complementation of split TEV and the problem is perhaps because of the dCas9 or gRNA.
+It was planned to use the ABI/PYL-splitTEV system as a control for the dCas9-splitTEV system, as stepping stones for troubleshooting which parts could be dysfunctional. For example, if the ABI/PYL system works but not the dCas9-splitTEV system, it is shown that the testing conditions can allow for complementation of split TEV, and the problem is perhaps because of the dCas9 or gRNA.