Ligations - help

Jane McHarg ejmcharg at exeter.ac.uk
Wed Jun 5 12:21:30 EST 1996


I have been trying to ligate a 3.1kb mutant pcr fragment into a 
homemade yeast/Ecoli shuttle expression vector unsuccessfully for 
weeks.  The vector has a Bam H1 site and is 7.5kb.  It is useful as it 
promotes my gene under its own promoter and as a result 50 -80% of the 
total cell protein is my gene product, so I must use this vector. 
Other people have successfully used this system.  The pcr frag is T7 - 
T3 and I then cut it with Bam H1 - I can confirm it has cut on a gel.
Help!



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