Oligo cloning

[Graham Atherton]

Try Linearising the plasmid, do not phosphatase, phenol extract.
Anneal oligo's in 100mM NaCl (80 degrees C down to RT, then leave on 
ice), Do not kinase.
Ligate 25-50ng vector with 1 microgramme annealed oligo 2-4 hrs RT.
Works every time for me (greater than 90% contain inserts) with added 
bonus that there are no multiple inserts.
Why solve problems if you can avoid them!
Graham Atherton
GRGGTA at UK.AC.MAN.CR.PICR