Team:Hong Kong-CUHK/DOC NBK MAY

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NOTEBOOK - MAY

1. Preparation of Competent Cells, aliquot the competent cells, stored in liquid nitrogen and then -80C refrigerator.
2. Transformation of: P123H, P53K, P421E


3. Mix content for N.P./ H.S. . Without the repaired casamino acid. RE analysis using XbaI and PstI, there are no expected bands were observed for CadC and   there are expected bands were observed for pSBIC
4. Mixing cultures for HS and NP
5. Make LB agar solution


6. Calibrate pH valve for N.P. and H.S. special medium
7. Preparation of medium for Natronomonas Pharaonis (N.P.) and Halobacterium Salinarum (H.S.). After autoclave, precipitate formed.
8. Re-mix the culture for N.P. and H.S. without autoclaving it. There was no precipitate formation and we successfully made the medium.


9. Mixing culture for H.S and N.P as the previous recipe calibration of pH
10. Checking of cultured N.P. and H.S. are viable or not by light microscope and there are no cells are found.
11. Make plate with antibiotics –Ampicillin.
12. Primer dilution of the 20 light ideas primers and sub-culturing of N.P. and H.S..
13. Amplification by using PCR, following by run gel of Sensory Rhodopsins Protein II gene.
14. Amplification by using PCR, following by run gel of Sensory Rhodopsins Protein I gene.


15. Amplification by using PCR, following by run gel of Htr I. Incorrect banding shown that the amplification using PCR fail.
16. Make plate with antibiotics-chloramphenicol.
17. Amplification by using PCR, following by run gel of Htr II. Correct banding shown and HtrI is being successfully amplified.
18. Amplification by using PCR, following by run gel of Tsr I. Correct banding shown and HtrI is being successfully amplified.


19. Amplification by using PCR, following by run gel of Tsr II. Incorrect banding shown that the amplification using PCR fail.
20. Redo PCR amplification of Htr I, following by run gel. Correct banding shown and HtrI is being successfully amplified.
21. Amplification by using PCR, following by run gel of Backbone Vector A. Incorrect banding shown.


22. Redo PCR amplification of Tsr II, following by run gel. Correct banding shown and HtrI is being successfully amplified.
23. Amplification by using PCR, following by run gel of Backbone Vector B. Incorrect banding shown that the PCR amplification was failed.

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