hello all,
i am hoping some of you might be able to help me. i am trying to run
sequencing reactions on a double stranded plasmid template. the problem is
that the plasmid is 56 kb. i have a protocol that has been successful for
plasmids up to twenty or twenty-five kb, but nothing for a plasmid this
large.
i ran some reactions today as follows:
template 2ug
primer 16pmol
i used a reaction volume of 40ul. the cycle sequencing conditions were
as follows:
95 for 5 minutes
forty cycles of
95 30 seconds
50 20 seconds
60 4 minutes
followed by a 4 degree hold.
these conditions gave no results.
If anyone thinks they can help, or has any suggestions please let me know.
I thank you all in advance,
-- gabriel archdeacon elim biopharmaceuticalsgabriel@elimbiopharm.com (650)-827-1984 fax (650)-827-1986
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