Required DNA concentrations for automated sequencing:

All samples and primers must be dissolved in distilled water, and must be at the standard concentrations listed below. We cannot do custom dilutions or reconstitute lyophilized samples.

WARNING! Primer concentrations are given in 'pMol/ul' - That's Picomoles per Microliter, NOT PICOMOLAR!

 

Template Type: Template Conc: Template Amt: Primer Conc: Primer Amt: Notes:
Bacterial Genomic DNA We have had some limited success with sequencing from complete bacterial genomic DNA. Ask the Director about this before submitting. 10 ul per reaction   per reaction Caution! Experimental! Talk to the Core Director.
PCR products (rule of thumb) For EACH kb of fragment size, 3 ng/ul 10 ul per reaction 1 pMol/ul 10 ul per reaction Important! Remove old PCR primers!!
Large DNA (plasmids over ~20 kb, cosmids, lambda clones) 500 ng/ul 10 ul per reaction 30 pMol/ul (30 uM) 20 ul per reaction Caution! Experimental! Talk to the Core Director.
DNA fragments (e.g. gel isolated fragments) For EACH kb of fragment size, 3 ng/ul 10 ul per reaction 1 pMol/ul 10 ul per reaction  
Single Stranded (e.g. M13) 25 ng/ul 10 ul per reaction 1 pMol/ul 10 ul per reaction  
Plasmids (mini-preps, maxi-preps, 4-15kb) 50 ng/ul 10 ul per reaction 1 pMol/ul 10 ul per reaction  
Large DNA (P1 clones, BACs, bacterial genomic) 1 ug/ul Yes, really! 10 ul per reaction 30 pMol/ul (30 uM) 20 ul per reaction Caution! Experimental! Talk to the Core Director.
siRNA Constructs 200 ng/ul 10 ul per reaction 3 pMol/ul 20 ul per reaction  
Rolling-Circle preps (e.g. 'TempliPHI' or equivalent) As per manufacturer's recommendation (typ 10X dilution of product) 10 ul per reaction 1 pMol/ul 10 ul per reaction  

Please re-check this table periodically whenever you submit samples, to ensure that we have not adjusted our recommendations.