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Team:Hawaii/Notebook/2008-09-13
From 2008.igem.org
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===PCR=== | ===PCR=== | ||
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:*oriT constuct from pSB1A2- for construction with J33207 for verification test | :*oriT constuct from pSB1A2- for construction with J33207 for verification test | ||
:*rep (pRL1383a)- to ligate to plac, B0030, and pSB1A3 and make a number of construct that can be sequenced with my new primers | :*rep (pRL1383a)- to ligate to plac, B0030, and pSB1A3 and make a number of construct that can be sequenced with my new primers | ||
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===Restriction Digest=== | ===Restriction Digest=== | ||
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| + | ::*b=from BB registry | ||
| + | ::*first #= colony from aada+pSB1A3 ligation (sequence verified) | ||
| + | ::*second#= colony from plac,B0034 ligation to the sequence verified aadA+pSB1A3 ligation | ||
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= Discussion = | = Discussion = | ||
Latest revision as of 08:13, 15 September 2008
| Projects | Events | Resources | ||
|---|---|---|---|---|
| Sponsors | Experiments | Milestones | Protocols | |
| Notebook (t) | Meetings (t) |
Things we did today
Wetlab work
Competent Cells
- Margaret, Krystle, Norman
- Today we made DH5-alpha competent cells. The first OD reading was between 0.3-0.4 for all three flasks. The OD after the last incubation with CMBD buffer was 1.1, 1.29, 0.8977, 1.079, and 0.942. (both of these OD readings are within the range we want.)
PCR
- Margaret
- oriT constuct from pSB1A2- for construction with J33207 for verification test
- rep (pRL1383a)- to ligate to plac, B0030, and pSB1A3 and make a number of construct that can be sequenced with my new primers
- omega interposon contruct from pSB1A2- to verify and sequence
- oriV(pRL1383a)- to ligate to pSB1A3 and sequence next week.
- (+) control pSB1A3 with VF2&VR
- (-) control water with VF2&VR
Restriction Digest
- Margaret
- Code
- a= aadA
- p=pRL1383a
- b=from BB registry
- first #= colony from aada+pSB1A3 ligation (sequence verified)
- second#= colony from plac,B0034 ligation to the sequence verified aadA+pSB1A3 ligation
| construct | RE-enzymes used | next step |
|---|---|---|
| J33207 | SP | dephosphorylate |
| ap36 | E, S | gel purify |
| ap32 | E, S | gel purify |
| ap61 | E, S | gel purify |
| ab95 | E, S | gel purify |
| ab92 | E, S | gel purify |
| ab96 | E, S | gel purify |
| ab94 | E, S | gel purify |
| ab98 | E, S | gel purify |
| ab93 | E, S | gel purify |
| base vector(I15020) | N | dephosphorylate |
| oriT(BB) | XP | ligate directly |
| oriT(my construct)PCR product 9/13 | XP | ligate directly |
| TT(B0016) | XP | ligate directly |
| pSB1A3 | EP | ligate to self(use as control) |
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 
]
