InsTA Clone t/a cloning blues( actually whites) - (Jul/25/2009 )
hi,
i am using insTA CLONE kit( fermentas). after ligation my insert(1.4 kb, gel puirified) into vector - the ligated product is showing increased size 3-4 bands! ( including correct one - faintest band)- after transformation and plating- 20ul and 50 ul in amp/iptg/xgal plates- i am only getting white colonies -no blue ones( altough colony obtained are proprotionate in control , 20ul and 50ul plates . colony pcr from various colonies is -ve!
help....
rabos on Jul 25 2009, 02:37 AM said:
i am using insTA CLONE kit( fermentas). after ligation my insert(1.4 kb, gel puirified) into vector - the ligated product is showing increased size 3-4 bands! ( including correct one - faintest band)- after transformation and plating- 20ul and 50 ul in amp/iptg/xgal plates- i am only getting white colonies -no blue ones( altough colony obtained are proprotionate in control , 20ul and 50ul plates . colony pcr from various colonies is -ve!
help....
Hi,
I am using INSTA cloning kit (fermentas) too. After colony PCR, the insert size is 2.5kb (1kb more than the supposedly 1.5kb.). Yeah,I only got white colonies. I guess I experienced the same thing as yours. Colony PCR of most of the white clones are negative. Really strange. Wonder what happen! PLs help us. Thanks..
Cheers,
appleyun
Dear frnds
I am cloning pcr product 450bp into TA Pcr2.1 3.9kb vector.......i am getting both blue and white colonies.....after colony PCR with M13 Forward + my primer reverse i am getting exact size 450bp for some samples.......for other set i am getting strong band of small size 250bp.......how come a nonspecific band is coming.....
please clarify me
thanks