Download Sequencing User Guide - ASU Shared Resources

Submitting Sequencing Samples
Purity and quantitation of the DNA template and primer dictate the quality of the sequencing
results. The templates should be purified and quantified before submitting for sequencing. We
can quantify templates for you, especially off campus customers; however this may slightly
delay the return of your data.
Optimal amounts for templates and primers are as follows:
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Single stranded DNA 100-200 ng
Double stranded DNA (plasmids) 150-200 ng
PCR products 10-30 ng (10ng for templates <500bp; 30ng for >1kb)
BAC or Cosmid DNA 500 ng
The amount of primer required for all of the above is 20 ng, except BAC and Cosmid which need
100 ng primer.
If you wish to set up reactions yourself, mix the template and primer at above amount and bring
the volume to 6 ul with dd H2O. For off campus customers we recommend drying the sample
prior to shipment to avoid possible loss, however properly sealed tubes can be successfully sent
too. If you prefer that we set up reactions for you, submit sufficient template and primer in
separate tubes.
Returning your Results
Sequencing
We routinely deliver data by email or dropbox.
Text files open with word or notepad. To open electropherogram files, you can use the programs
at the following links:
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Sequence scanner software from ABI
Finch TV
Fragment analysis
We can deliver data by dropbox and if you need software to view chromatograms go here:
 ABI free software
Sequencing Primers
The following primers can be added by the DNA Lab.
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T3 5’CAA TTA ACC CTC ACT AAA GGG 3’
T7 5’ TAA TAC GAC TCA CTA TAG GG 3’
SP6 5’TAT TTA GGT GAC ACT ATA G 3’
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SP6* 5’GAT TTA GGT GAC ACT ATA G 3’
M13F 5’ GTT GTA AAA CGA CGG CCA GT 3’
M13R 5’AGG AAA CAG CTA TGA CCA TG 3’
M13R (–23) 5’AGC GGA TAA CAA TTT CAC ACA GG’
T7 terminator 5’ GCT AGT TAT TGC TCA GCG G 3’
BGH-R 5’ TAG AAG GCA CAG TCG AGG C 3’