Team:BYU Provo/Team Thermosensor/Week11

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==Week 11 (June 26 - Jul 2)==
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===6/27/11===
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 +
It looks like 33-1 worked, but I don’t know if lacZ and GFP were actually inserted
 +
 +
Doesn’t look like lacZ did, but GFP looks promising.
 +
 +
===6/28/11===
 +
 +
PCR purfication of 33-1 (lacZ PCR)
 +
 +
* Qiaquick PCR purification kit protocol -Julie
 +
 +
lacZ restricition digest (TRD 34-1)
 +
 +
* T33-1 PCR up/lacZ with 1 ul of HindIII and XbaI each
 +
 +
pPLAT-Bad restriciton digest (TRD 34-2)
 +
 +
* pIG 13 with 1 ul each of HindIII and XbaI
 +
 +
Ran a low melt. Cut out pPLAT-Bad and stuck in 1.5 centrifuge tube. Could not see lacZ band...->doesn’t matter, I used wrong lacZ reverse
 +
 +
We could run another low melt and we may want to PCR it again. Other half of low melt in fridge.
 +
 +
lacZ PCR (35-1)
 +
 +
* Phusion polymerase and 1 ul each of IG40 and IG41 primers with pIGGY template
 +
* Making two in case there is another purification problem or something
 +
 +
35-2: PCR up of lacZ according to qiaquick instructions
 +
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===6/30/11===
 +
 +
Running gel on lacZ PCR products
 +
 +
* Both lanes look good. Now onto restriction digest.
 +
 +
RD lacZ (35-3)
 +
* 1-2 ul each restricition enzyme (HindIII and XbaI)
 +
 +
35-4 Ran low melt and cut out lacZ band. Very faint, but there!
 +
 +
lacZ/pPlatbad ligation 36-1
 +
 +
* T4 ligase, 3 ul vector, 3 ul lacZ (added a little extra b/c faintness of band)
 +
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===7/1/11===
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 +
lacZ/pPlatbad transformation
 +
 +
Followed Grose protocol
 +
 +
Plated 100 ul on LB-amp and incubated overnight
 +
 +
Labeled T36-1 lacZ trans 1 July 2011 - Devin
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Latest revision as of 08:26, 24 September 2011

Team BYU Provo

BYU Provo
 

Contents

Week 11 (June 26 - Jul 2)

6/27/11

It looks like 33-1 worked, but I don’t know if lacZ and GFP were actually inserted

Doesn’t look like lacZ did, but GFP looks promising.

6/28/11

PCR purfication of 33-1 (lacZ PCR)

  • Qiaquick PCR purification kit protocol -Julie

lacZ restricition digest (TRD 34-1)

  • T33-1 PCR up/lacZ with 1 ul of HindIII and XbaI each

pPLAT-Bad restriciton digest (TRD 34-2)

  • pIG 13 with 1 ul each of HindIII and XbaI

Ran a low melt. Cut out pPLAT-Bad and stuck in 1.5 centrifuge tube. Could not see lacZ band...->doesn’t matter, I used wrong lacZ reverse

We could run another low melt and we may want to PCR it again. Other half of low melt in fridge.

lacZ PCR (35-1)

  • Phusion polymerase and 1 ul each of IG40 and IG41 primers with pIGGY template
  • Making two in case there is another purification problem or something

35-2: PCR up of lacZ according to qiaquick instructions

6/30/11

Running gel on lacZ PCR products

  • Both lanes look good. Now onto restriction digest.

RD lacZ (35-3)

  • 1-2 ul each restricition enzyme (HindIII and XbaI)

35-4 Ran low melt and cut out lacZ band. Very faint, but there!

lacZ/pPlatbad ligation 36-1

  • T4 ligase, 3 ul vector, 3 ul lacZ (added a little extra b/c faintness of band)

7/1/11

lacZ/pPlatbad transformation

Followed Grose protocol

Plated 100 ul on LB-amp and incubated overnight

Labeled T36-1 lacZ trans 1 July 2011 - Devin

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