Difference between revisions of "Part:BBa K1463000:Design"
(→Design Notes) |
(→Design Notes) |
||
| Line 11: | Line 11: | ||
Using both HindIII and PstI produces different sized fragments dependent on the orientation of the switch promoter.<br><br> | Using both HindIII and PstI produces different sized fragments dependent on the orientation of the switch promoter.<br><br> | ||
Transcriptional terminators were designed into the switch construct in order to prevent transcriptional leakage of GFP when the switch is in the "OFF" position; this also applies to prevent RFP transcription in the "ON" position.<br><br> | Transcriptional terminators were designed into the switch construct in order to prevent transcriptional leakage of GFP when the switch is in the "OFF" position; this also applies to prevent RFP transcription in the "ON" position.<br><br> | ||
| − | A spacer was included in the switch design to ensure the att sites were suitable distance apart for the recombinase enzyme to work without steric hindrance. | + | A spacer was included in the switch design to ensure the att sites were a suitable distance apart for the recombinase enzyme to work without steric hindrance. |
===Source=== | ===Source=== | ||
Latest revision as of 15:57, 17 October 2014
Recombinase Switch With GFP and RFP
- 10COMPATIBLE WITH RFC[10]
- 12INCOMPATIBLE WITH RFC[12]Illegal NheI site found at 813
Illegal NheI site found at 836 - 21INCOMPATIBLE WITH RFC[21]Illegal BamHI site found at 793
- 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal AgeI site found at 34
Illegal AgeI site found at 146 - 1000INCOMPATIBLE WITH RFC[1000]Illegal BsaI.rc site found at 711
Illegal BsaI.rc site found at 1668
Design Notes
Restriction sites in switch - HindIII
HindIII was designed into the switch downstream of the promoter. This allowed for confirmation of the "flip" of the switch by restriction digest.
Using both HindIII and PstI produces different sized fragments dependent on the orientation of the switch promoter.
Transcriptional terminators were designed into the switch construct in order to prevent transcriptional leakage of GFP when the switch is in the "OFF" position; this also applies to prevent RFP transcription in the "ON" position.
A spacer was included in the switch design to ensure the att sites were a suitable distance apart for the recombinase enzyme to work without steric hindrance.
Source
Switch synthesised by IDT from a given sequence, RFP and GFP are from the 2014 distribution.