IGEM:UNAM LCG/2009/Notebook/Hydrobium etli/2011/09/24: Difference between revisions

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(Autocreate 2011/09/24 Entry for IGEM:UNAM_LCG/2009/Notebook/Hydrobium_etli)
 
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==Entry title==
==HydG ligation==
* Insert your content here.


===Abstract===
Now that we have HydG digested and purified we need to put it into the vector pSB1C3 to send it to the iGEM.
I first ran a gel to see HydG's concentration and then I calculate the ammounts of pSB1C3 to make the ligation. Unfortunatly it looks like I lost a lot of HydG during the band extraction so the ligation was done with very little ammounts of DNA.
===Gel Details===
I ran this gel to know HydG's concentration.
*120V, 40min.
*5μL HydG + 2μL dye; 2μL ladder
====Gel image====
Lanes:
  1.HydG
  2.Ladder [http://www.fermentas.com/en/products/all/dna-electrophoresis/ogeneruler-dna-ladders/sm117-ogeneruler-mix fermentas]
  The rest of the lines are not of interest for this analysis.
[[Image:HydG_DD_aislado_concentracion.JPG]]
====Gel analysis====
The concentration is VERY low, the mark at 1700bp aprox is barely visible. [[User:Gustavo_A._Ruiz_Buendia|Gustavo]] helped me to calculate the concentration, we decided that it is 0.003pM
===HydG-pSB1C3 ligation===
We need to do this ligation to send HydG it to the registry
====Concentration calculation====
*HydG = 0.0006 pM/μL
*pSB1C3 = 0.012 pM/μL
*Relation: 0.6:12
Since we need a 3:1 cassete:vector relation we want a 0.2 vector concentration to get 0.6:0.2
*Final volume and final concentration: 18[0.2]
*Needed volume and actual concentration: X[0.12]




Revision as of 22:33, 24 September 2011

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HydG ligation

Abstract

Now that we have HydG digested and purified we need to put it into the vector pSB1C3 to send it to the iGEM.

I first ran a gel to see HydG's concentration and then I calculate the ammounts of pSB1C3 to make the ligation. Unfortunatly it looks like I lost a lot of HydG during the band extraction so the ligation was done with very little ammounts of DNA.

Gel Details

I ran this gel to know HydG's concentration.

  • 120V, 40min.
  • 5μL HydG + 2μL dye; 2μL ladder

Gel image

Lanes: 

 1.HydG
 2.Ladder fermentas
 The rest of the lines are not of interest for this analysis.

Gel analysis

The concentration is VERY low, the mark at 1700bp aprox is barely visible. Gustavo helped me to calculate the concentration, we decided that it is 0.003pM


HydG-pSB1C3 ligation

We need to do this ligation to send HydG it to the registry

Concentration calculation

  • HydG = 0.0006 pM/μL
  • pSB1C3 = 0.012 pM/μL
  • Relation: 0.6:12

Since we need a 3:1 cassete:vector relation we want a 0.2 vector concentration to get 0.6:0.2

  • Final volume and final concentration: 18[0.2]
  • Needed volume and actual concentration: X[0.12]



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