Interrupted reading frame, help!
genecutl at mendel.berkeley.edu
Wed Apr 13 23:38:43 EST 1994
In article <1994Apr13.134355.1 at wehi.edu.au>, chai_z at wehi.edu.au wrote:
> Hello there,
> I'm cloning an insulin or insulin-realated genes from a mollusc. I pulled out
> a cDNA using oligo probe of a reference sequences. My sequence showed 87%
> homology with the reference sequence, but by taking away 4 bases in the middle.
> The extra bases interrupted the reading frame. Furthermore, the whole cDNA
> (1488 bases) contained many stop codons, which resulted in very short reading
> frames (the biggest one is about 70 aa).
> Any comments are appreciated.
> Zhonglin Chai
> (chai_z at wehi.edu.au)
Two possibilities come to mind. The insertion may be a cDNA arifact.
Reverse transcriptase has been known to make mistakes. A more likely
possiblity, considering the abundance of stops is that you've cloned a
pseudogene. There are examples of pseudogenes that can still be transcibed
but not translated. I would advise using this cDNA to do a Southern to
see if there are multiple related genes. Also, I'd take this clone and
use it to screen a genomic library. The genomic library wouldn't be
affected by the cloning artifacts that can screw up cDNA libraries.
More information about the Methods