Sequencing AT rich Templates

[Buzby, Philip]

When nucleotide mixtures in sequencing kits are developed, the key
assumption is templates will have an nearly equal ratio of A:G:C:T's.  The
dNTP/dye-ddNTPs are then optimized with this in mind.  When one sequences
templates with a predominance of A/T or G/C, the fact is there is not enough
of that base(s) in the mixture to permit long sequences.  Not having access
to the terminators is a real problem, but you might try adding/spiking in
additional unlabeled dATP/TTP for A/T rich or dGTP/dCTP for G/C templates.
Of course this upsets the dNTP/ddNTP ratio so that longer products will now
predominate.

Philip R. Buzby, Ph.D.

---