Team:Hawaii/Notebook/2008-08-17
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===Verification of transformants=== | ===Verification of transformants=== | ||
:<strong> Grace</strong> | :<strong> Grace</strong> | ||
- | [[Image:081708colonyPCR.jpg|right|thumb| | + | [[Image:081708colonyPCR.jpg|right|thumb|350px|EtBr stained 2.5% agarose gel ran at 95V for 1 .5 hours. Five microliters of PCR reaction were loaded into each well.]] |
{| class=wikitable border=1 align=center | {| class=wikitable border=1 align=center | ||
! DNA | ! DNA | ||
Line 12: | Line 12: | ||
! Blue colonies | ! Blue colonies | ||
|- | |- | ||
- | | align=center|BB-pRL1383a + J33207 | + | | align=center|BB-pRL1383a <br>+ J33207 |
- | | align=center|LB + sp<sub>100</sub> + sm<sub>50</sub> + IPTG (1mM) + X-gal (2mM) | + | | align=center|LB + sp<sub>100</sub> + sm<sub>50</sub> +<br> IPTG (1mM) + X-gal (2mM) |
- | |align=center| | + | |align=center|105 |
- | |align=center| | + | |align=center|0 |
|- | |- | ||
- | | align=center|BB-pRL1383a + J33207 | + | | align=center|BB-pRL1383a <br>+ J33207 |
|align=center|LB + sp<sub>100</sub> + X-gal (2mM) | |align=center|LB + sp<sub>100</sub> + X-gal (2mM) | ||
- | |align=center| | + | |align=center|44 |
- | |align=center| | + | |align=center|0 |
|- | |- | ||
- | |align=center| BB-pRL1383a (negative control) | + | |align=center| BB-pRL1383a <br>(negative control) |
- | |align=center|LB + sp<sub>100</sub> + sm<sub>50</sub> + IPTG (1mM) + X-gal (2mM) | + | |align=center|LB + sp<sub>100</sub> + sm<sub>50</sub> +<br> IPTG (1mM) + X-gal (2mM) |
- | |align=center| | + | |align=center|44 |
- | |align=center| | + | |align=center|0 |
|- | |- | ||
- | | align=center|no DNA (negative control) | + | | align=center|no DNA <br>(negative control) |
- | |align=center|LB + sp<sub>100</sub> + sm<sub>50</sub> + IPTG (1mM) + X-gal (2mM) | + | |align=center|LB + sp<sub>100</sub> + sm<sub>50</sub> +<br> IPTG (1mM) + X-gal (2mM) |
- | |align=center| | + | |align=center|0 |
- | |align=center| | + | |align=center|0 |
|} | |} | ||
+ | :* X-gal did not work? | ||
:* Colony PCR to verify insert | :* Colony PCR to verify insert | ||
+ | ::* ~320bp band is due to pRL1383a plasmid | ||
+ | ::* BB-pRL1383a + J33207 bands are smaller than (-) control bands, potential success in replacing GFP w/ lac device | ||
+ | :::* (-) bands due to contamination | ||
+ | :* Restreaked colonies to purify | ||
+ | :* Grew up colonies in TB+sp<sub>100</sub> liquid media for plasmid prep on Tuesday | ||
= Discussion = | = Discussion = |
Latest revision as of 05:32, 18 August 2008
Projects | Events | Resources | ||
---|---|---|---|---|
Sponsors | Experiments | Milestones | Protocols | |
Notebook (t) | Meetings (t) |
Things we did today
Wetlab work
Verification of transformants
- Grace
DNA | Plate | Total colony forming units | Blue colonies |
---|---|---|---|
BB-pRL1383a + J33207 | LB + sp100 + sm50 + IPTG (1mM) + X-gal (2mM) | 105 | 0 |
BB-pRL1383a + J33207 | LB + sp100 + X-gal (2mM) | 44 | 0 |
BB-pRL1383a (negative control) | LB + sp100 + sm50 + IPTG (1mM) + X-gal (2mM) | 44 | 0 |
no DNA (negative control) | LB + sp100 + sm50 + IPTG (1mM) + X-gal (2mM) | 0 | 0 |
- X-gal did not work?
- Colony PCR to verify insert
- ~320bp band is due to pRL1383a plasmid
- BB-pRL1383a + J33207 bands are smaller than (-) control bands, potential success in replacing GFP w/ lac device
- (-) bands due to contamination
- Restreaked colonies to purify
- Grew up colonies in TB+sp100 liquid media for plasmid prep on Tuesday
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 ]