From
P. Lalitha
Centre for Biotechnology
Anna University
Madras 600 025
Tel 91 44 2350298
Fax 91 44 2350299
email : biospic at giasmd01.vsnl.net.in
I am working with the cDNA library of W. bancrofti
constructed in lambda Zap II vector. While screening the
library with filarial antibodies we came across a
particular phage which lysis the E.coli strain XL1 Blue
within four hours. The titre of the phage is extremely high
(around 10exp12). Since this particular phage reacts very
strongly with the antisera we tried to characterise this
phage.
USING VARIOUS STRAINS OF E.COLI:
With KL-226 it does not form plaques. This confirms that it
is not a T1 phage.
With Q358 and LE392 strain it forms plaques this confirms
it is a lambda phage.
With CSH57() strain it doesn't form plaques. This confirms
that it is a Lytic lambda.
DNA EXPERIMENTS:
The phage DNA was extracted according to the prescribed
protocols as in Mannitias et al 1989. The size of the Phage
DNA was confirmed by running it in a 0.5% agarose gel using
the wild type lambda and the DNA marker. Next we tried to
see the restriction pattern of the DNA using the Hind III
enzyme for linearisation and Xho1/Xba1 for double digest.
But we did not see any restriction pattern and only streaks
were visible.
CONVERSATION INTO PLASMID:
Since the titre of the Phage is very high we were not able
to convert it into plasmid using helper phage .
I NEED THE FOLLOWING HELP
1. Why streaks appear when digested with restriction
enzymes?.
2. Why this phage lysis within 4hrs and what is the reason
for very high titre?
Can any one help me regarding this phage.
