Team:PKU Beijing/Notebook/AND Gate 1/Core/SupD

Notebook > AND Gate 1 > Core > Shan Shen's Note

2009.7.23
11:30 Recycle all the inserts of SupD and terminator.

12:20 Digest high-copy bi-stable plasmids.

12:30 Start to digest.

12:40 Ligation of SupD+terminator and 1-18A (strongest promoter).

16:50 Electrophoresis to recycle the high-copy bi-stable digestion products. The order and the amount of the samples: control plasmids 2μL, digestion products, 20μL marker 5μL.

2009.7.24


10:30 There are six clones on the plate. Shake those clones in the incubator. PCR clones to test if they are correct. The results are not very good.

14:00 Transform the plasmids PKD46 from France in two tubes.

22:40 Miniprep 1-18A + SupD + terminator.

2009.7.28
00:20 Digest 1-18A + SupD +terminator plasmids to test if they are correct. Digest them into vectors as well. Digest into inserts: Digest into vectors: PCR at the same time:

1:00 Start to digest.

1:30 Start to PCR.

12:00 Electrophoresis to test if the PCR products and digestion products are correct. The order and the of the samples: marker, PCR products 1, PCR 2, PCR 3, PCR4, PCR5, PCR6, digestion control plasmids, digestion1, digestion2, digestion3, digestion4, digestion5, digestion 6.

Results:

16:00 Transfrom pKD3, pcp 20 to DH5a. Digest 1-18C into vectors.

20:00 Electrophoresis the digestion products: The order and the amount of the samples: marker 10μL, plasmids control 2μL, digestion 1 15μL, digestion2 15μL. Results: The digestion products are correct.

2009.7.29
10:00 Recycle the digestion products. Miniprep the plasmids of 1-18C and PYFP.

13:30 Digest plasmids PYFP from France.

13:48 Start to digest.

15:00 Transform psp20 into DH5a. Ligation of 1-18C backbone and SupD+ terminator inserts.

15:30 PCR to standardize the PYFP plasmids. Take a gradient from 54 centigrade to 58 centigrade.

17:30 Digest the high-copy bi-stable PCR products.

19:30 Electrophoresis the digestion products and PCR products. The order of the samples: marker, plasmids control, digestion products, PCR products under 54 centigrade, PCR products under 56 centigrade, PCR products under 58 centigrade. Results: Digestion products are weird, while the PCR products under 58 centigrade are specialized. As a result, recycle the PCR products under 58 centigrade.

21:55 Digest PYFP plasmids.