Part:BBa_K929302
AID in Potsdam Standard
AID in Potsdam Standard
Introduction
| AID in Potsdam Standard | |
|---|---|
| |
| BioBrick Nr. | BBa_K929302 |
| RFC standard | [http://2012.igem.org/Team:Potsdam_Bioware/Project/Potsdam_Standard RFC 10], [http://2012.igem.org/Team:Potsdam_Bioware/Project/Potsdam_Standard Potsdam Standard] |
| Requirement | pSB1C3 |
| Source | existing part:(BBa_K103001) |
| Submitted by | [http://2012.igem.org/Team:Potsdam_Bioware Potsdam_Bioware2012] |
AID is known to be responsible for somatic hypermutation and the class-switch recombination of immunoglobulin in B cells. This enzyme of 28 kDa originally occurs in B cells but does also show activity after transfection into CHO cells. AID induces the deamination of cytidine to uridine at actively transcribed single strand DNA. The replacement of cytidine by uridine leads to a mismatch during DNA replication and integrates a single base substitution predominantly in the immunoglobulin genes.
Cloning with Potsdam Standard
To generate this part, we used the [http://2012.igem.org/Team:Potsdam_Bioware/Project/Potsdam_Standard Potsdam Standard] for cloning the AID into the Potsdam Standard Backbone. Here, we tested four different ligation conditions: 1. with electrophoretic unpurified, digested Potsdam Standard backbone with and without ligase and 2. with electrophoretic purified digested Potsdam Standard backbone with and without ligase. To compute the efficiency, we counted the colonies without and with red fluorescence indicating a failed ligation reaction (fig. 1). The results are summarized in fig. 2. We can see that with ligase the ligation success is much higher than without, and that a electrophoretic purification of the backbone improve the ligation efficiency.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000INCOMPATIBLE WITH RFC[1000]Illegal BsaI.rc site found at 84
Illegal SapI site found at 185
| None |