IGEM:Peking/2007/Count-Conjugation-Notebook/2007-8-11: Difference between revisions
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==after ligation & mini-prep== | ==after ligation & mini-prep== | ||
NEXT DAY(8-12): | |||
*From ''left'' to ''right'': | |||
1,4,5 PlacI-I741051 @ EcoRI/PstI | |||
2,3,4 J61003 @ EcoRI/PstI | |||
7 I741051 @ EcoRI/PstI | |||
[[Image:Peking_2007-8-12_1_5_6J61003E-p_2_3_4-plac-I74051E-P_8-I74051E-P.jpg]] | |||
=oriT Knock Out= | =oriT Knock Out= |
Revision as of 08:35, 13 August 2007
Tandem-OriT by Qu Mingzhi
OUR FIRST Biobricks !!
- F-OriT-J23066-OriT pSB1A2 now named as I741051!
- waiting for sequencing to get the final detail.
re-do: PlacI -> I741051
- basic experiment condition :IGEM:Peking/2007/Count-Conjugation-Notebook/2007-8-10
- we changed "fast T4 ligase" ligation time : from 20min ->10 min @16℃
- digesting for longer time.
- use both "fast T4 ligase" & normal T4 ligase".
electrophoresis result
I741051 vector
- From left to right:
- Maker (DL2000 Plus)
- I741051 vector (after Gel extraction, EcoRI/XbaI)
R0010 before gel extraction
- From left to right:
- R0010 -1 @ EcoRI/SpeI
- R0010 -2 @ EcoRI/SpeI
- Maker (DL2000 Plus)
R0010 after gel extraction
- R0010
- Maker (DL2000 Plus)
after ligation & mini-prep
NEXT DAY(8-12):
- From left to right:
1,4,5 PlacI-I741051 @ EcoRI/PstI 2,3,4 J61003 @ EcoRI/PstI 7 I741051 @ EcoRI/PstI
oriT Knock Out
- By Xu Anting
Transformation of pKO3-oriT-deleted plasmid into DH5a
- Result: negative controls have an approximately number of clones in comparing with ligation products. It seems that our DH5a have been contaminated.
Ligation Products Verification
- By Colony PCR
- Picked colonies and grew them in an additional Cm Petri-dish, in 37 centrigrade, overnight.