Almost any program for work with DNA sequences will do some oligo
design. One very popular and exclusively focused on oligo design
software product is "Oligo" (for DOS, Mac and Windows), it will search
for the best primers, and will give you annealing temperatures, loop &
dimer formation. Their Web site is at http://www.oligo.net/ whereby
you can download a free demo (the real version is not for free). My
suggestion is that "Oligo" should be your first choice. It has voice
capabilities.
My favorite is, however, "Prophet" (for Windows), because it is
"all-in-one" software that takes HUGE sequences (e.g. 600000 bases),
which other programs may stumble upon, cuts them with virtually any
restriction enzyme (and shows the results in circular or linear graphs
or sequence with cutting sites), translates in all six frames and shows
molecular weights, allows multiple listings (and saving) of different
sequences in the same window, does comparisons (alignments), searches
for primers etc. I like a lot the "Protein-to-DNA" reverse translation
that lets you input various codon usage tables for different species.
In addition, Prophet will do statistical analysis (any, not only
DNA-related). "Prophet" was developed by BBN Technologies under the
sponsorship of the NIH and was FREE (!!!) last year (I do not know about
current situation). Last year the Web download was for free, but it was
necessary to register afterwards (again for free), I do not know whether
this year they changed the policy (or disappeared from the radar
screen). Some universities bought a license and can re-distribute
including ftp (see http://www.basic.nwu.edu/biotools/prophet.html).
Disadvantages of the Prophet: no voice reading, no loops and dimers
viewing after the primer selection, although self-complementarity is a
parameter with variables that can be modified during the initial step of
criteria setting for the primer search.
Another "all-in-one" program that will do (arguably) a better primer
design and that does have a voice is MacMolly Tetra (for Apple-Macintosh
computers), a German software (not for free).
And last, but not least, we did a small survey and found out that most
of our clients use the good old "eye" method of selecting primers, with
a couple of simple basic rules (GC content of 50% or so, GC-rich 5'-end
and AT-rich 3'-end, 18-23 bases, sequence comparison against the GenBank
etc.) that ensure a decent selection. Anyway, even the most carefully
selected oligo may end up annealing not only to your gene of interest,
but to yet unknown pseudogene or another region in the 3 Gb human
genome, if your template is complex. At the other hand, an
"incorrectly" chosen oligo may still work very well, especially if the
template is, say, a purified plasmid. An awful 20-mer, in which the
first 7 bases are the reverse complement of the last 7, can still do a
fine job in PCR and ThermoSequenase reactions at annealing temperatures
of 60 degrees and higher, because at those temperatures the loops and
dimers will not form.
Best regards,
victor, Alpha DNA
www.alphadna.com
_____________________________________
Chris Jacobs wrote:
I need feedback on the best/worst programs for designing PCR and
sequencing
primers. (both Mac and PC programs).
Thanks!
Chris Jacobs
DNA Sequencing Lab
Parke-Davis Pharmaceuticals
Office Phone: (734) 622-5724
Lab Phone: (734) 622-1815
E-mail: chris.jacobs@wl.com