Question on reverse transcriptase(s)

[Ashok Aiyar]

Hello: 
 
I have a question/survey about reverse transcriptases. 
 
There are two enzymes that are commercially available, the MuLV 
 and AMV reverse transcriptases (RTs). 
 
The differences between the two are as follows: 
 
	AMV					MuLV 
 
a) 2 subunits				1 subunit 
b) active 45 degrees			active 42 degrees 
c) optimum temp = 42 degrees		optimum temp = 37 degrees 
d) contains intrinsic 			no endonuclease activity 
     endonuclease  
e) No RNase H mutant is			RNase mutant  is 
     commercially available		sold commercially 
f) Purified from virions		Bacterially produced 
g) The AMV enzyme is supposed to have greater processivity than 
	the MuLV enzyme 
 
As part of my Ph.D, I am in the process of cloning the two subunits of 
the avian enzyme (AMV RT).  I am also making some mutations in the 
enzyme, including one that knocks out the intrinsic endonuclease. 
 
Although there is no reason for me to do this, I could also fairly 
easily make one that knocks out the RNase H activity, without 
affecting processivity.  If I made this mutation, I would be interested 
in making it freely available. 
 
My question is: 
 
"Given your choice of the following 4 enzymes for a cDNA synthesis, 
	or primer extension experiment, which would you use?" 
 
a) Native MuLV RT (no endonuclease, possesses RNase H) 
b) Altered MuLV RT (no endonuclease, RNase H knocked out) 
c) Native AMV RT (intrinsic endo, possesses RNase H) 
d) Altered AMV RT (no endonuclease, RNase H knocked out) 
 
Please mail your response and comments to axa12 at po.cwru.edu 
 
Thank you, 
 
Ashok Aiyar 
Graduate Student in Biochemistry 
Department of Biochemistry 
CWRU Medical School 
Cleveland, OH 44106-4935 
 
Disclaimer: 
 
My views (and deeds) are my own, and Case Western Reserve University  
is not responsible for any of them. 
-- 
Ashok Aiyar
axa12 at po.cwru.edu
aiyar at cwbio.bioc.cwru.edu