Y. Sugino et M. Morita, A NEW DNA CLONING SEQUENCING VECTOR WITH A BUILT-IN MECHANISM FOR GENERATION OF NESTED DELETIONS USING TRANSPOSON TN3/, Gene, 148(1), 1994, pp. 169-170
We have constructed a new cloning/sequencing vector suitable for seque
ncing of DNA fragments too long to be sequenced in a single step. This
vector plasmid contains inverted repeats (IR) of transposon Tn3, the
kil and c1857 genes of phage lambda, and multiple cloning sites (MCS).
Escherichia coli cells harboring plasmids containing nested deletions
from one of the Tn3 IR ends, across kil, to variable end points, can
be positively selected by plating at 42 degrees C. The deletion produc
ts, fractionated according to their size by agarose-gel electrophoresi
s, can be sequenced by using a synthetic primer whose 3'-end is locate
d within the Tn3 IR, and the total sequence of the insert can be const
ructed from the partial sequences.