Team:WHU-China/Notebook

• Dealing with biobricks
• Preparation of competent cells (Using Calcium Chloride)
• Plasmid Extraction（Using Plasmid Mini Kit）
• Transformation (heat-shock method)
• Cleavage with two restriction enzymes
• Get Extraction
• Identification of DNA by Electrophoresis
• Ligation
• PCR of the colony
• Identification of DNA by Sequencing
• Others............

The forth layer assembly and assembled CHL1-3:
8/26 extract plasmid: 009-13,009-3. Enzyme digest 009-13,009- 3. Result shows that it is not cut completely .PCR 20 + 2 positive control.Transformation: 2 M-13 J-2 M-20 F, PSB, 2 K-16 MIV, CHL1-CHL3 (DH5 α and TrpR defects strains)
8/27 extract plasmid: 8H, 2M-14 H-2M. enzyme digest 009-3, 8H, 2 M-14 H-2 M. Ligate 009-2-3.Identification of the transformation of PCR
8/26 ligation results. Cut CHL1 religate CHL1-CHL3, CHL1-CHL4.Enzyme cut PCB, 23 L, RLS1.Transformation: CHL1-CHL3 (DH5 α and TrpR defects plant), CHL1-CHL4 (DH5 α and TrpR defects plant), 009-2-3, controls.
8/28 repeat 27th enzymes digest,ligated 009-2-3. Transformation: 18 M-RLS1, 18 O-RLS1, 2 G Ⅱ-RLS1, PSB-23 L, 009-2-3, CHL1-CHL3 (TrpR defects plant), CHL1-CHL4 (TrpR defects strains) controls. Reserve RLSF-2, PSB-1.....

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