From 2011.igem.org

Revision as of 20:28, 26 July 2011

Gels for DspB

2011.07.03-2011.07.09

Figure 1: Gel proving the successful transformation of WT dspB from UBC into E. coli Top10. Lane 1: ladder; lanes 2-4: digests (EcoRI and PstI) of miniprep DNA from overnights of transformant colonies 1-3. There is a clear band at the appropriate size for dspB in all of the experimental lanes.	Figure 2: Gel of transformants that should contain both WT dspB and a promoter insert. Lane 1: ladder; lanes 2-4: WT dspB with PrsaA colonies 1-4; lanes 5-7: WT dspB with Pxyl colonies 1-4; lane 8: standard dspB with no promoter.	Figure 3: Follow up to Figure 2. Gel showing WT dspB with various promoter inserts as contrasted to a standard dspB with no promoter. Lane 1: WT dspB + PrsaA colony 3; lane 2: with Pxyl colony 3; lanes 3,4: with BBa_K081005 colonies 3 and 4; lane 5: standard WT dspB; lane 6: ladder.	Figure 4: Test of more WT dspB with Pxyl colonies. Lane 1: ladder; lanes 2-5: WT dspB with Pxyl colonies 5-8; lane 6: standard dspB with no promoter.

2011.07.10-2011.07.16

Figure 5: Optimized dspB transformed into pSB1C3 colonies A-E. Lane 1: ladder; lanes 2-5: optimized dspB colonies A-E; lanes 6-9: optimized esp. Colony D shows the expected size.	Gel of colony PCR of transformants that should carry WT dspB behind the Pxyl promoter. Lanes 1-4: colonies 9, 10, 12, and 15 from transformation plate; lane 5: dspB WT standard from PCR.	Gel for extraction of WT dspB behind Pxyl. Lane 1: ladder; lanes 2,3: miniprep digest.	Gel of colony PCR of transformants carrying optimized dspB and rsaA C-terminal. Lane 1: ladder; lanes 2-5: transformation colonies 1-4; lane 6: dspB standard from PCR of WT dspB.
Lane 1: ladder; lanes 2-5: transformants that should carry PrsaA, optimized dspB, and rsaA; lanes 6-9: transformants that should carry Pxyl, optimized dspB, and rsaA; lane 10: standard optimized dspB from PCR. None of these transformants showed success.	Lane 1: ladder; lanes 2-5: transformants that should carry Pxyl, optimized dspB, and rsaA; lane 6: standard optimized dspB with rsaA from PCR. None of these colonies were successful.	Lane 1: standard optimized dspB with rsaA from PCR; lanes 2,3: transformants that should carry WT dspB behind Pxyl; lane 4: ladder. No successful transformations.	Lane 1: ladder; lanes 2-5: transformants that should carry BBa_K081005, optimized dspB, and rsaA; lane 6: standard optimized dspB with rsaA fragment. Lanes 3 and 5 carry the desired combination of genes.
Lane 1: ladder; lanes 2-7: transformants that should carry Pxyl, optimized dspB, and rsaA; lane 8: standard fragment of optimized dspB with rsaA. None of the transformants carried the desired gene sequence.

2011.07.17-2011.07.23

Lane 1: ladder; lanes 2-5: transformants that should carry BBa_K081005, optimized dspB, and rsaA in pMR10; lanes 6-9: transformants that should carry PrsaA, optimized dspB, and rsaA in pMR10; lane 10: standard DNA fragment of optimized dspB with rsaA. Lane 7 carries the desired gene sequence.	Gel of PCR of transformants that should carry Pxyl, optimized dspB, and rsaA. Lane 1: ladder; lanes 2-6: PCR products; lane 7: standard fragment of dspB with rsaA.	Gel of PCR of transformants that should carry Pxyl, optimized dspB, and rsaA. Lane 1: ladder; lanes 2-9: transformant PCR products; lane 10: digested miniprep of what should have been WT dspB, but showed up as the wrong fragment size.	PCR of transformants that should carry BBa_K081005, optimized dspB, and rsaA in pMR10. Lane 1: ladder; lanes 2-7: PCR products.
PCR of transformants that should carry Pxyl, optimized dspB, and rsaA in pMR10. Lane 1: ladder; lanes 2-8: PCR products.