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From 2010.igem.org

Contents 1 Andreas 1.1 Cloning and assembly 1.1.1 Transformation results 1.1.2 Colony PCR 1.1.3 Gel verification 1.2 Sequencing

Andreas

Cloning and assembly

Transformation results

From 25/9 transformations

Good colony yield on all plates. Good white:red colony ratio on pEX plates (Constructs 1, 2, 3 and 4).

Colony PCR

Picked colonies for colony PCR.

1. pEX.N-LMWP⋅SOD⋅His (K): 1-2
2. pEX.N-TAT⋅SOD⋅His: 1-2
3. pEX.N-Tra10⋅SOD⋅His: 1-2
4. pEX.N-LMWP⋅SOD⋅His (C): 1-2
5. pSB1K3.N-LMWP⋅SOD⋅His.RBS.yCCS: 1-4
6. pSB1K3.N-TAT⋅SOD⋅His.RBS.yCCS: 1-4
7. pSB1K3.N-Tra10⋅SOD⋅His.RBS.yCCS: 1-4
8. pSB1C3.N-LMWP⋅SOD⋅His.RBS.yCCS: 1-4
9. BL21 pEX.N-TAT⋅SOD⋅His 3: 1-2
10. BL21 pEX.N-TAT⋅SOD⋅His 4: 1-2

Standard colony PCR protocol.

• Elongation time: 2:00

Gel verification

Gel 1
1 % agarose, 110 V

Gel 2
0.8 % agarose, 90 V

Expected bands

1. 744 bp
2. 735 bp
3. 765 bp
4. 744 bp
5. 1645 bp
6. 1636 bp
7. 1666 bp
8. 1645 bp
9. 735 bp
10. 735 bp

Results
Gels run too long. New gels will be run tomorrow.

Sequencing

Samples prepared 25/9 were sent for sequencing. Sequencing information added to the 25/9 notebook page.