Posted 17 August 2011 - 11:58 PM
I have ligated my insert to the PENTR /D TOPO vector and trying hard to get my insert band after enzyme digestion.. Its really surprising for me that when i digest my plasmid with ASCI and NOt I enzyme , i cannot see the positive band for my insert but the vector size that is 2580 bp looks ok , my pro is that if i digest the same plasmid with different enzyme like ASCI and Xbai ,in that case i got my required insert but the vector size become very high may be more than 5000 bp... i tried this many time and got same result now i am stuck????? What to do i have no idea ??
if some body have same experience plz do share with me or any suggestion ...
Pcr product size 1530 bp
Vector size 2580 bp
waiting for reply
Posted 18 August 2011 - 04:49 AM
Posted 18 August 2011 - 05:06 PM
Posted 18 August 2011 - 07:43 PM
Better, though, would be to find a clone with the right properties.
Posted 19 August 2011 - 04:48 AM
Posted 21 August 2011 - 03:33 AM
Anyways thakns for your suggestion
Posted 22 August 2011 - 04:07 PM
i tried with single enzyme digestion but the size of plasmid is very big .
my required size is around 4000 bp as 2580 vector + 1530 insert but the result is around 7000 bp???
i dont know whats going wrong do you have any idea ???
Posted 22 August 2011 - 05:24 PM