2009.6.9 (with LinMin)
11:10
The e-coli cultivated yesterday did not grow
The material of the former culture medium went bad. Need renewing.
11:29
Practise minipreping the plasmids
12:58
Practise digesting the plasmids
2009.6.13 (with LinMin)
15:00
make the culture medium, repair computer, search Parts:
| Parts Code | Location | Location | Plasmid | Containment | Description
|
| BBa_E0240 | Kit Plate 1 | 12M | Psb1A2 | GFP rbs-repoter | Medium rbs
|
| BBa_E0840 | Kit Plate 1 | 12O | Psb1A2 | GFP rbs-repoter | Strong rbs
|
| BBa_R0080 | Kit Plate 1 | 12E | Psb1A2 | Ara Promoter |
|
| BBa_C0062 | Kit Plate 1 | 4O | Psb1A2 | Lux R gene | No LVA
|
| BBa_C0051 | Kit Plate 1 | 4E | Psb1A2 | C1 repressor | With LVA
|
| BBa_C0179 | Kit Plate 2 | 8M | Psb1A2 | lasR activator | No LVA
|
| BBa_C0079 | Kit Plate 1 | 14J | Psb1A2 | lasR activator | With LVA
|
| BBa_R0079 | Kit Plate 1 | 12A | Psb1A2 | LasR/Pal promoter |
|
16:55
dissolve the plasmids, prepare to transform
add 15uL ddH2O
21:20
Transform plasmids above
2009.6.14
13:10
repair the computer
14:08
make the LB culture medium
21:00
search the rbs parts:
| Parts Code | Location | Location | Plasmid | Containment | Description
|
| BBa_B0030 | Kit Plate 1 | 1H | Psb1A2 | RBS | strong rbs
|
| BBa_B0031 | Kit Plate 1 | 2G | Psb1A2 | RBS | Weak
|
| BBa_B0032 | Kit Plate 1 | 2I | Psb1A2 | RBS | Medium
|
| BBa_B0033 | Kit Plate 1 | 2K | Psb1A2 | RBS | Weaker
|
| BBa_B0034 | Kit Plate 1 | 2M | Psb1A2 | RBS |
|
| BBa_J44001 | Kit Plate 1 | 1J | Psb1A2 | RBS |
|
| BBa_J61100 | Kit Plate 1 | 5J | Psb1A2 | RBS |
|
| BBa_J61101 | Kit Plate 1 | 5L | Psb1A2 | RBS |
|
| BBa_J61107 | Kit Plate 1 | 5N | Psb1A2 | RBS |
|
| BBa_J61117 | Kit Plate 1 | 11L | Psb1A2 | RBS |
|
| BBa_J61127 | Kit Plate 1 | 11N | Psb1A2 | RBS |
|
2009.6.15
21:07
Transform plasmids above (2009/06/14)
21:48
begin to transforming
2009.6.16
11:50
cultivation is finished The results come out well
2009.6.19 (with Wu Shuke)
21:11
Select clones from the plate
2009.6.20
23:00
transform 1-12L
2009.6.21
9:50
Transform the 2008 Parts’ RBS
11:54
finish transformaion
2009.6.22
9:14
Transformation is failed, redoing
22:30
re-transformation
2009.6.23
1:00
start the cultivation
13:00
the second transformation is failed
2009.6.24
14:59
Prepare to make TOP10 Sensitive Cells
16:00
Begin to make LB, etc
17:30
Search for LuxR s
| BBa_R0062 | 09 Kit1 | 6O | pSB1A2 | Promoter
|
| BBa_R0065 | 09 Kit1 | 8C | pSB1A2 | Promoter
|
| BBa_R1062 | 09 Kit1 | 8G | pSB1A2 | Promoter
|
23:38
Cultivate TOP10 Cells
2009.6.25
21:44
Transform the rbs:
1003-12C; 1003-12G; 1004-2A; 1004-2C; 1004-3G
2009.6.26
22:36
Cultivate:
1004-1C; 1004-3C; 1004-4C; 1004-2G
23:07
Prepare to make 1-2I Sensitive Cells
23:15
Cultivate 1-2I Cells
2009.6.27
15:58
Miniprep the plasmids:
1004-1C; 1004-3C; 1004-4C; 1004-2G Failed
16:09
Make the 1-2I Sensitive Cells Failed
2009.6.28
20:18
Miniprep the plasmids:
1004-1C; 1004-2C; 1004-3C; 1004-4C; 1004-2G
21:37
Make the 1-2I Sensitive Cells
2009.6.29
17:40
Check the 1-2I Sensitive Cells Huge Success! The protocol is added to ftp
20:09
Prepare to make DH5A Sensitive Cells
2009.7.3
18:00
Search Parts:
| BBa_J37032 | 1003 | 6C | pSB1A2 | GFP controlled by LuxRP
|
22:44
Cultivate 1003-6C Failed. The plasmid is bad.
2009.7.3-2009.7.10
Searching for papers on T3P and T3Pol, which is basic for the second logic gate. Contact with the writer to get the plasmids.
During this time, a review on T7 and T3 systems is summarized, which has been sent to ftp.
2009.7.6
19:51
Miniprep the plasmids:
| 2-4O | LuxR+RBS | Amp+
|
| 1-18P | P(cat) | Kan+
|
21:08
Digest the plasmids of 2-4O and 1-18P:
| Digestion System Insert | 20μL
|
| Plasmids | 5μL
|
| Pst1 | 1μL
|
| Xbal1 | 1ul
|
| Buffer 1*M | 2μL
|
| dd H2O | 11μL
|
| Digestion System Vector | 20μL
|
| Plasmids | 5μL
|
| Pst1 | 1μL
|
| Spe1 | 1ul
|
| Buffer 1*H | 2μL
|
| dd H2O | 11μL
|
2009.7.7
0:34
The digestion reaction begins
7:46
add 0.4uL cip and 2ul buffer to the vector system
8:07
the result come out as follows:
18:03
Recycle the fragments.
19:40
Ligate the 2-4O insert to 1-18P vector:
| Insert | 3μL
|
| Vector | 1μL
|
| Buffer 10* | 1μL
|
| Ligase | 1uL
|
| dd H2O | 4μL
|
19:48
Ligation start
23:44
Begin the transformation of the ligation plasmid
2009.7.8
16:57
Cultivate the 2-4O->1-18P
2009.7.9
9:06
Miniprep the 2-4O->1-18P plasmids
11:52
Digest the plasmids of 2-4O->1-18P and 1-23L:
| Digestion System Insert | 20μL
|
| Plasmids | 5μL
|
| EcoR1 | 1μL
|
| Spe1 | 1ul
|
| Buffer 1*H | 2μL
|
| dd H2O | 11μL
|
| Digestion System Vector | 20μL
|
| Plasmids | 5μL
|
| EcoR1 | 1μL
|
| Xbal1 | 1ul
|
| Buffer 1*M | 2μL
|
| dd H2O | 11μL
|
12:13
Reaction Start
17:32
Stop the digestion
20:08
Prepare to Ligation
20:17
Ligation start
|
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