Difference between revisions of "Part:BBa K3505023"
(→Design Notes) |
(→Verification of Cloning) |
||
Line 19: | Line 19: | ||
===Verification of Cloning=== | ===Verification of Cloning=== | ||
− | [[File:T--Thessaly--FFAR2-ECFP-digestion.png|700px|thumb|none|<i><b>Fig. | + | [[File:T--Thessaly--FFAR2-ECFP-digestion.png|700px|thumb|none|<i><b>Fig.3:</b>(U=Uncut C=Cut) Restriction Digestion of FFAR2:V2tail:TCS with BamHI, Expected bands : 3204 bp, Positive Result:C2,C3</i>]] |
===Source=== | ===Source=== |
Latest revision as of 00:28, 28 October 2020
FFAR2:V2tail:TCS GB compatible with B2-B3
Usage and Biology
FFAR2 is a naturally found eukaryotic GPCR protein that exhibits high affinity for SCFAs (acetic, propionic and butyric acid)(Kaemmerer, 2010) . After background searching through the NCBI database, we have identified the gene coding sequences needed for the designing of this gene construct. More specifically, a vasopressin receptor 2 segment has been attached to the C-terminal of the receptor, as it mediates recruitment of a TEV tagged b-arrestin-2, along with a TEV cleavage site(TCS).
Design Notes
The coding sequence was domesticated . We removed BsmBI ,BsaI , BtgZI, BpiI sites in order to be compatible with GoldenBraid and MoClo. The sequence is presents in pUPD2 and has overhangs compatible for GoldenBraid cloning. The CDS has position B2-B5.
Verification of Cloning
Source
Synthesized by IDT.
Sequence and Features
- 10INCOMPATIBLE WITH RFC[10]Illegal PstI site found at 284
Illegal PstI site found at 864 - 12INCOMPATIBLE WITH RFC[12]Illegal PstI site found at 284
Illegal PstI site found at 864 - 21INCOMPATIBLE WITH RFC[21]Illegal BamHI site found at 106
- 23INCOMPATIBLE WITH RFC[23]Illegal PstI site found at 284
Illegal PstI site found at 864 - 25INCOMPATIBLE WITH RFC[25]Illegal PstI site found at 284
Illegal PstI site found at 864
Illegal NgoMIV site found at 364 - 1000INCOMPATIBLE WITH RFC[1000]Illegal SapI site found at 560
Illegal SapI site found at 815
Illegal SapI.rc site found at 17
References
- Kaemmerer, E., 2010. Fatty acid binding receptors in intestinal physiology and pathophysiology. World Journal of Gastrointestinal Pathophysiology, 1(5), p.147.