genepool

Next Generation Sequencing and Bioinformatics
at the University of Edinburgh


Methods used by the GenePool

Sanger Sequencing: BigDye Controls Protocol.

For BigDye Terminator Cycle Sequencing ready Reaction Kit.

Quarter reactions

                                1 sample    1 plate 
   5X Sequencing Buffer         2 µl        212 µl
   ddH20                        1 µl        106 µl 
   M13 Primer (0.8pmol/ µl)     4 µl        424 µl
   BigDye                       2 µl        212 µl 
   PGEM (0.2ug/ µl)             1 µl        106 µl

Eighth reactions

                                 1 sample    1 plate 
   5X Sequencing Buffer          2 µl        212 µl
   ddH20                         1 µl        212 µl 
   M13 Primer (0.8pmol/ µl)      4 µl        424 µl
   BigDye                        2 µl        106 µl 
   PGEM (0.2ug/ µl)              1 µl        106 µl
 

132 µl in first strip , transfer 10 µl to each well

Slow cycle

   95°C 30sec 
   50°C 20sec 
   60°C 4min

The GenePool has core support from: GenePool admin site
edinburgh
The University of Edinburgh
School of Biological Sciences
The Darwin Trust
of Edinburgh
darwin
mrc
nerc
bbsrc
sulsa

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This is version 2.2 of the GenePool website (July 2009).