09/07/ 24 (yy/mm/dd) Experimenter Kashima, Takashima, Hibino, Miyazawa Note-taker Takashima Title Flow Chart Sample Date Sample NameEnzymepurification KitProductStorage iGEM DNA parts kits 3-O-2 3-A-4 3-O-2 3-A-4 EcoR1 EcoR1, PST1 7/24 Miniprep Kit, Restriction Enzyme Digestion Electrophoresis 1.Miniprep Kit 2. Restriction Enzyme Digestion 3.Electrophoresis Restriction Enzyme Digestion 6F freezer

Miniprep Kit (Discussion) Cultured sample ( :30 ～ :50 ) ;3-O-2,3-A-4 Centrifuge 4 ℃, × g, 2min Remove the top of layer twice P1 250ul P2 250ul N3 350ul Centrifuge 4 ℃, × g, 10min Apply the top of layer to column Centrifuge RT, 10sec and remove through Buffer PB 500ul Centrifuge RT, 10sec and remove through Buffer PE 650ul Centrifuge 4 ℃, × g, 1min Exchange tube (remove C 2 H 5 OH) Milli Q 50ul RT 1min Centrifuge 4 ℃, × g, 1min Collect through and measure concentration

Result of Miniprep Kit (Discussion) Sample:conc. (ng/ul) 3-O A

Description ( Electrophoresis ) (Discussion) Gel :Agarose Time : 12:50 ～ 13:20 Voltage: 100V 1234 Sample 3-O-23-A-43-O-23-A-4 3ul enzyme EcoR1 0.17ul 0.43ul enzyme H2OH2O1ul 1.4ul H buffer8.2ul 8.1ul Total12.7ul 12.9ul PST1 0.3ul samplename3-O-23-A-43-O-23-A-4Marker volume ×lodding buffer ℃ incubate ( time: :50 ～ :20 )

Result of Electrophoresis (Discussion) This result indicates that the parts ( 3-O-2, 3-A-4) were treated with enzyme in appropriate way. 1000bp 5000bp １ 234M 3-O-23-A-4 parts length(bp) plasmid length(bp) 3266