Team:SPSingapore/Notebook-Week-8

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  • Week 1 (24/5 - 30/5)
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Research Notebook

Week 8 (12/7 - 18/7) ▪ July 13 Anaerobic Promoter    ☀ Yi Han & Yun Ting ☀ Ligation for FNRgfp (4X) 200ng of gfp plasmid (4.36ul) 297.5 ng insert (7:1) 10.3 1ul 10Buffer (53.34) 4ul T4 ligase Trial run of anaerobe chamber open sachet to decrease O2 at 3.30pm takes 2.5h to activate streak plate of P. putida a strict aerobe and E. coli, facultative aerobe in chamber at 30deg C to grow O/N P. putida is an obligate aerobe and if chamber works ,it will not grow E. coli should grow in both conditions controls grown outside chamber ▪ July 14 Anaerobic Promoter    Invasin + Listerolysin ☀ Yi Han ☀ No colonies for FNRgfp plasmid Inv Plasmid Verification buffer 1 EcoRI 0.5 in plasmid 0.5 dH2O 8 Total 10ul Gel extraction RE digest of EsaR-GFP plasmid Ligation Control Vector (45ng) 1 1 insert 1 1 Buffer 1 1 dH2O 6 7 ligase 1 1 Transformation of FNRgfp Anaerobe jar E. coli grew in both conditions P putida-> some growth in anaerobe chamber Proper streak plate in aerobic conditions Packet runs out after 16 hours ▪ July 15 Anaerobic Promoter    ☀ Yi Han ☀ FNRgfp-> no colonies after transformation religation (4X reaction) 200ng of gfp plasmid (4.4ul) 2125ng of insert (8ul) 4ul T4 ligase 55.6 H2O Anaerobic Promoter    ☀ Yi Han & Chi Yan ☀ Transformation of FNR gfp into 20ul of BL21 ▪ July 16 Anaerobic Promoter    ☀ Yi Han ☀ No colonies grew Ran a gel, FNRgfp pcr, gfp pcr, fnr, 100bp ladder ▶ Seems to be band shift.