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From 2012.igem.org

Week 24: 8 october - 14 october

CCMV

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12 October

Transformed IPTG_CCMV_NEG into a BL21 strain. Transformed biobrick RFP in pSB1C3 and the one with IPTG + RBS in kanamycin backbone into DH5A.

14 October

Inoculated 10 mL LB with a colony from each transformation plate.

GFP modification

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11 October

• grow JM109 containing our biobricks BBa_K883702 (GFP coil with an IPTG induced promoter) and BBa_K883703 (GFP coil + His tag with an IPTG induced promoter)and induce expression with IPTG

-> no conclusion could be made due to growth on the negative control

• send samples of BBa_K883702 BBa_K883703 BBa_K883701 and BBa_K883700 in for sequencing to check if the last transformation from pJET into the iGEM backbones succeded

-> sequencing results where never obtained - the samples did not arrive

12 October

• 2nd time growing JM109 containing our biobricks BBa_K883702 (GFP coil with an IPTG induced promoter) and BBa_K883703 (GFP coil + His tag with an IPTG induced promoter) and induce expression with IPTG

-> again there is growth on the negative control -> no green fluorescence could be seen

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PLRV

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We started another production process to obtain PLRV VLPs. We considered that production of the CCMV VLP with the negative inside had worked well in the first batch which produced with the well acknowledged production strain BL-21, and not in the later batch from the JM-109 strain. Because our last attempt to produce PLRV VLPs was done in JM-109 cells as well, we figure it may be possible that the production strain was the problem. Therefore, we will try the same procedure again but now in BL-21 E. coli.

We transformed electrocompetent BL-21 cells with Bba_K883401, which contains a PLRV Coat Protein encoding gene under the IPTG inducible promotor.

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