need help,why colony pcr of my blue has product what i need - (Nov/20/2005 )
i need to pcr 18s rdna to make phylogenetic relation .but i did colony pcr to detect my work,i found that the blue clone sitll have my target band,around 2000bp(18s+part of the t-vector )although the band is weak,not so strong.except the band of 2000 bp, my white clone still have another band,which is about 3000bp around. the vector i use is T-vector,i use TA clone.
i wondered that is it t-vector self-ligation,but why the blue have my target band .
i am a chinese grduate student,my english is poor,please forgive me .
this picture is my gel.
the marker is 4500,3000,2000,1200,800,500,200.
number 9,10 is blue clone.the others is white clone.
Blue/white selection is imperfect in many ways. All it tests is whether the beta-lactamase alpha complementation protein is being made and is functional. If your insert is in-frame and not too big, or if it happens to have a leader which is good enough to initiate translation and plausible amino acid sequences, the protein can be made even with an insert. If you are having trouble finding a colony with insertions, it is always a good idea to pick a few blue colonies (as you have discovered). Be happy.
thank you ,phage434.i am very happy that somebody gave me reply.