Virginia Commonwealth/23 July 2009

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(Difference between revisions)
(→Thursday 23 July 2009)
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** pick controls as well to make more seed and electrocompotent stock
** pick controls as well to make more seed and electrocompotent stock
[[User:Trentay|Trentay]] 17:25, 23 July 2009 (UTC)
[[User:Trentay|Trentay]] 17:25, 23 July 2009 (UTC)
 +
''Kevin and Adam''
 +
*We need to PCR DNA padding onto the first nine synthesized promoters.  When we ordered these oligo's we did not consider that we needed additional overhang from the BioBrick prefix and suffix in order for the restriction enzymes to work properly.  Doing this PCR will correct that error. [[User:Bussingkm|Bussingkm]] 20:56, 23 July 2009 (UTC)
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====Wetlab====
====Wetlab====
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* Cryogenic stocks were made
* Cryogenic stocks were made
[[User:Trentay|Trentay]] 17:25, 23 July 2009 (UTC)
[[User:Trentay|Trentay]] 17:25, 23 July 2009 (UTC)
 +
''Kevin and Adam''
 +
*First we reconstitute the DNA.  The rule of thumb is that you take 1/2 the amount of oligo's (nmol) of H2O in uL for ~100x DNA stock solution.  Store in (-20C)
 +
**A problem was encountered while reconstituting the DNA.  For several of the sequences the amount of water described above was not sufficient to reconstitute the DNA.  It was too viscus and could not be pipetted at all.  We doubled the amount of water that was used and made note of the approximate concentration.
 +
 +
{| cellpadding="10" border="1" alignment="center"
 +
|##|| ~nmol DNA ||uL H2O || Concentration
 +
|-
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|Design 1 || 25 nmol || 12.5 uL || 100x
 +
|-
 +
|Design 2 || 25 nmol || 25 uL || 50x
 +
|-
 +
|Design 3 || 25 nmol || 25 uL || 50x
 +
|-
 +
|Design 4 || 25 nmol || 12.5 uL || 100x
 +
|-
 +
|Design 5 || 25 nmol || 25 uL || 50x
 +
|-
 +
|Design 6 || 25 nmol || 25 uL || 50x
 +
|-
 +
|Design 7 || 25 nmol || 25 uL || 50x
 +
|-
 +
|Design 8 || 25 nmol || 25 uL || 50x
 +
|-
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|Design 9 || 25 nmol || 25 uL || 50x
 +
|-
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|Forward end padding|| 48.3 nmol || 24.15 uL || 100x
 +
|-
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|Reverse end padding || 36.4 nmol || 18.2 uL || 100x
 +
|}
 +
|}

Revision as of 20:56, 23 July 2009

Contents

Thursday 23 July 2009

Results

Maria and Afton

  • All overnight cultures grew except one vial of NEB 10 beta seed stock
    • there was a second vial of seed stock that did grow successfully
  • Transformations were successful
Name Type Plate Purpose Growth Observation
NEB 10 beta seed stock LB make more seed stock lawn
NEB 10 beta electro comp. LB (+) Control/ make more stock lawn
NEB 10 beta electro comp. LB (+) Control shocked lawn
DB 3.1 electro comp. LB (+) Control shocked lawn
DB 3.1 electro comp. LB (+) Control/make more stock lawn
NEB 10 beta electro comp. Cm (-) Control no growth
NEB 10 beta electro comp. Cm J23100 w/ J06702 6 small colonies

Trentay 17:19, 23 July 2009 (UTC)

Tasks

Maria and Afton

  • Make cryogenic stocks of all overnight cultures
  • Make stocks of parts I1352 (RFP) and I13522 (GFP)
  • Update notebook, documents
  • Pick colonies from transformation
    • pick controls as well to make more seed and electrocompotent stock

Trentay 17:25, 23 July 2009 (UTC) Kevin and Adam

  • We need to PCR DNA padding onto the first nine synthesized promoters. When we ordered these oligo's we did not consider that we needed additional overhang from the BioBrick prefix and suffix in order for the restriction enzymes to work properly. Doing this PCR will correct that error. Bussingkm 20:56, 23 July 2009 (UTC)

Wetlab

Maria and Afton

  • Cryogenic stocks were made

Trentay 17:25, 23 July 2009 (UTC) Kevin and Adam

  • First we reconstitute the DNA. The rule of thumb is that you take 1/2 the amount of oligo's (nmol) of H2O in uL for ~100x DNA stock solution. Store in (-20C)
    • A problem was encountered while reconstituting the DNA. For several of the sequences the amount of water described above was not sufficient to reconstitute the DNA. It was too viscus and could not be pipetted at all. We doubled the amount of water that was used and made note of the approximate concentration.
## ~nmol DNA uL H2O Concentration
Design 1 25 nmol 12.5 uL 100x
Design 2 25 nmol 25 uL 50x
Design 3 25 nmol 25 uL 50x
Design 4 25 nmol 12.5 uL 100x
Design 5 25 nmol 25 uL 50x
Design 6 25 nmol 25 uL 50x
Design 7 25 nmol 25 uL 50x
Design 8 25 nmol 25 uL 50x
Design 9 25 nmol 25 uL 50x
Forward end padding 48.3 nmol 24.15 uL 100x
Reverse end padding 36.4 nmol 18.2 uL 100x

|}

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