Worm Breeder's Gazette 11(2): 78

These abstracts should not be cited in bibliographies. Material contained herein should be treated as personal communication and should be cited as such only with the consent of the author.

Further Studies on Tc1 Transcription and Translation

A.D. Radice, N.E. Sherif and S.W. Emmons

In order to understand the mechanisms underlying Tc1 excision and 
transposition, we are studying transcription and translation of the 
Tc1 open reading frame.  To identify the 5' end of a possible Tc1 
message, we have performed primer extension experiments with polyA+ 
RNA from various strains as the template.  Using three primers located 
at various sites within the Tc1 ORF, we have identified a number of 
potential 5' ends.  These 5' ends result in primer extension products 
that are distinct from the products of premature termination of 
reverse transcriptase seen when sense RNA synthesized in vitro is used 
as the template.  To gain further evidence for transcripts initiating 
within Tc1, we carried out PCR amplification experiments after first 
strand cDNA synthesis from polyA+ RNA preparations.  We wanted to know 
whether there was a region of Tc1 that could not be amplified in this 
way, suggesting the presence of a 5' end.  So far the longest region 
we have amplified spans from nucleotide 304 to 1548 (putative 
polyadenylation signal).  Our inability so far to amplify with two 
different upstream primers in the region adjacent to the inverted 
repeat (nucleotides 60 through 100), and a downstream primer at 
position 924-943, suggests that the 5' end may be located in the 
region between nucleotide 60 and nucleotide 304, and that no read 
through transcripts extend past position 943.
As reported at the Cold Spring Harbor meeting in May, we have three 
antisera that recognize TcA.  Two were raised against synthetic 
peptides, and one against a TrpE-TcA fusion protein.  After 
purification by binding to antigen, these antisera reacted primarily 
with a single protein of 34-36 kd in nematode homogenates from various 
strains.  We have now found that this protein has a higher molecular 
weight than the 273 aa TcA protein synthesized in E.  coli (kindly 
provided by R.  Plasterk).  This result is similar to the one reported 
by R.  Plasterk at the meeting.  Therefore, the protein in nematodes 
either has additional amino acids, or is otherwise modified, or it is 
an unrelated, crossreacting protein.