Difference between revisions of "Part:BBa K4421029"

Revision as of 12:25, 16 July 2022

NFAT_RE-Gal4-KRAB

Gal4-KRAB(BBa_K2446037) transcription inhibitor was constructed under the control of the NFAT-RE promoter(BBa_K4421021).

Special Note

To simplify the transfection process and improve the transfection efficiency, we decided to transfect pSV40-UAS-iCASP9-2A-GFP and NFAT_RE-Gal4-KRAB on the same plasmid. But the KRAB protein has been demonstrated to be capable of inhibiting all promoters within at least 3 kB and the forward construct would therefore inhibit the NFAT-RE promoter, we generated the opposite construct, in which the two expression cassettes were cloned in such a manner that both promoters were at opposite ends and at a long distance.

The structure of NFAT_RE-Gal4-KRAB

Sequence and Features

Assembly Compatibility:

10
INCOMPATIBLE WITH RFC[10]
Illegal PstI site found at 478
12
INCOMPATIBLE WITH RFC[12]
Illegal PstI site found at 478
21
COMPATIBLE WITH RFC[21]
23
INCOMPATIBLE WITH RFC[23]
Illegal PstI site found at 478
25
INCOMPATIBLE WITH RFC[25]
Illegal PstI site found at 478
Illegal NgoMIV site found at 263
Illegal NgoMIV site found at 398
1000
COMPATIBLE WITH RFC[1000]

@@ Line 4: / Line 4: @@
 Gal4-KRAB(<partinfo>BBa_K2446037</partinfo>) transcription inhibitor was constructed under the control of the NFAT-RE promoter(<partinfo>BBa_K4421021</partinfo>).
+===Special Note===
+To simplify the transfection process and improve the transfection efficiency, we decided to transfect pSV40-UAS-iCASP9-2A-GFP and NFAT_RE-Gal4-KRAB on the same plasmid. But the KRAB protein has been demonstrated to be capable of inhibiting all promoters within at least 3 kB and the forward construct would therefore inhibit the NFAT-RE promoter, we generated the opposite construct, in which the two expression cassettes were cloned in such a manner that both promoters were at opposite ends and at a long distance.
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