"
Team:Hawaii/Notebook/2008-10-17
From 2008.igem.org
(Difference between revisions)
(→Restreaked BBpRL1383a construct) |
(margaret) |
||
| Line 12: | Line 12: | ||
:* Inoculated 200 ml TB+sp<sub>2.5</sub>+sm<sub>2.5</sub> with blue colony. Incubated over the weekend at 37C with shaking at 250 rpm. | :* Inoculated 200 ml TB+sp<sub>2.5</sub>+sm<sub>2.5</sub> with blue colony. Incubated over the weekend at 37C with shaking at 250 rpm. | ||
| + | |||
| + | ===PCR verification of constructs=== | ||
| + | :<strong>Margaret</strong> | ||
| + | |||
| + | :*20 colonies from each of [[Team:Hawaii/Notebook/2008-10-16|yesterday's]] transformation were verified by PCR today using the VF2 and VR primers. | ||
| + | |||
| + | |||
| + | == Drylab work == | ||
| + | ===Sequencing=== | ||
| + | :<strong> Margaret</strong> | ||
| + | |||
| + | :*Results for plac/rbs(30)/rep(7) came in. The promoter and rbs were not present. The only discrepancies in the sequence were due to bad sequence and could be explained by overlapping sequences. | ||
| + | :*This construct is already ligated to oriV and the oriT/aada so next step will be to isolate that plasmid, digest with E,S and ligate the promoter and rbs in front. rbs (B0034) will be used because we have primers for this rbs and it can be easily verified. | ||
| + | |||
= Discussion = | = Discussion = | ||
Revision as of 03:16, 19 October 2008
| Projects | Events | Resources | ||
|---|---|---|---|---|
| Sponsors | Experiments | Milestones | Protocols | |
| Notebook (t) | Meetings (t) |
Things we did today
Wetlab work
Restreaked BBpRL1383a construct
- Grace
- Looked at J33207+pRL1383a (BBpRL1383a) plates. 50 μl plate had blue colonies. Restreaked on LB+sp2.5+sm2.5 + X-gal
Inoculated for plasmid prep
- Grace
- Inoculated 200 ml TB+sp2.5+sm2.5 with blue colony. Incubated over the weekend at 37C with shaking at 250 rpm.
PCR verification of constructs
- Margaret
- 20 colonies from each of yesterday's transformation were verified by PCR today using the VF2 and VR primers.
Drylab work
Sequencing
- Margaret
- Results for plac/rbs(30)/rep(7) came in. The promoter and rbs were not present. The only discrepancies in the sequence were due to bad sequence and could be explained by overlapping sequences.
- This construct is already ligated to oriV and the oriT/aada so next step will be to isolate that plasmid, digest with E,S and ligate the promoter and rbs in front. rbs (B0034) will be used because we have primers for this rbs and it can be easily verified.
Discussion
Quote of the Day
History is the only laboratory we have in which to test the consequences of thought. - Étienne Gilson
[http://manoa.hawaii.edu/ 
][http://manoa.hawaii.edu/ovcrge/ 
][http://www.ctahr.hawaii.edu 
]
