Team:SPSingapore/Notebook-Week-6

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• NOTEBOOK
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• PROTOCOL
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• ENTRIES
  • Week 1 (24/5 - 30/5)
  • Week 2 (31/5 - 6/6)
  • Week 3 (7/6 - 13/6)
  • Week 4 (14/6 - 20/6)
  • Week 5 (21/6 - 27/6)
  • Week 6 (28/6 - 4/7)
  • Week 7 (5/7 - 11/7)
  • Week 8 (12/7 - 18/7)
  • Week 9 (19/7 - 25/7)
  • Week 10 (26/7 - 1/8)
  • Week 11 (2/8 - 8/8)
  • Week 12 (9/8 - 15/8)
  • Week 13 (16/8 - 22/8)
  • Week 14 (23/8 - 29/8)
  • Week 15 (30/8 - 5/9)
  • Week 16 (6/9 - 12/9)
  • Week 17 (13/9 - 18/9)

Research Notebook

Week 6 (28/6 - 4/7) ▪ June 28 esa Quorum Sensing    ☀ Adrian ☀ BBPrefix_esaRBS PCR H2O 221ul Buffer 80ul MgCl1 32ul dNTP 32ul FP_Bioricks Prefix 16ul ORP_esaRBS_fragsyn 16ul synpart_BBP_esaRBS 1ul gotaq 2ul Total 400ul ▶ Success-> PCR purification esaRBS_GFP_BBsuffix H2O 237ul Buffer 80ul MgCl2 32ul dNTP 32ul RP_BB_suffix 16ul ORP_esaRS_GFP 16ul plac_GFPPLASMID 1ul gotaq 2ul ▶ Failed -> troubleshooting-> new OFP_esaRBBS_GFP     Redid 30/6 with new primers     Success with new primers-> fusion pcr ▪ June 29 Lab Safety    ☀ SPSingapore Team ☀ Safety Inspection for our lab by OSHE. We passed with flying colours! ▪ July 2 esa Quorum Sensing    ☀ Clarice ☀ Fusion PCR with esaRBS, GFP H2O 104.07ul Buffer 40ul MgCl2 16ul dNTPs 16ul FP_BB_prefix 8ul RP_BB_prefix 8ul 400ng esaRBS 2.13ul 400ng esaRBSgfp 4.8ul gotaq 1ul total 200ul ▶ annealing temp 50degC ▶ Gel electrophoresis: looks correct-> PCR Purification RE digest of esaRBS+GFP insert Buffer 2 EcoRI/PstI 1/1 Template 2ug 5ul dH2O 11ul Total 20ul RE digest of esaRBS+GFP vector Buffer 2ul EcoRI/PstI 1/1 Template 4ul 2ug 5ul dH2O 12ul Total 20ul ▪ July 3 Anaerobic Promoter    ☀ Yi Han ☀ Ran gel of gfp plasmid EcoRI/PstI, took out gel slice for vector - 4kb ▶ gel extraction ▶ gfpvector->38.7ng/ul ▶ fragment-> direction purification 226.4ng/ul Ligation reaction (5:1) 6X 300ng vector 7.8ng/ul 531.2ng of insert 2.4ul 12ul buffer 6ul enz 91.8ul H2O ▶ ligate for 2 hours at 16 hours ▶ transform, plate, grow ON ▪ July 4 Anaerobic Promoter    ☀ Yi Han ☀ No colonies-> ligation did not work Recalculate ligation reaction (3:1),7x 350ng vector->9ul vector 446.3ng insert-> 2ul 7ul ligase 110 H2O 12ul buffer ▶ ligate for 2 hours at 16 hours ▶ transform, plate, grow ON