ligation problem

Fabien Jovelin f.jovelin at eurosequence.com
Tue Jul 24 08:53:50 EST 2001


Dear all,
I have to ligate two digested fragment with cohesive ends. These fragments
are 2.7 and 6 kb long. Ligation is made at 14°C using Sigma kit in a total
volume of 60 µl. Running results on agarose gel shows a degradation pattern
with 3 very "poor" bands corresponding to the fragments and the expected
ligated fragment; my problem is that these band of interest is too faint to
be purified, and located into a degraded lane...
How can I improve my ligation yield ? Any help welcome.
PS : ligase is fine because it works in other experiments...
Is the high final volume a problem ?

Fabien

--
Fabien Jovelin
ESGS-Cybergène






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