Hi all,
I have a question here which I can't seem to figure out the answer.
I have a cell line with an integrated retrovirus. Each cell has only 1 copy per cell and I specifically know where the integration site is. Using 2 pairs of primer - the primer pair is made up of 1 in the host and 1 in the respective (5' or 3' LTR) - I was able to amplify and confirm the integration site.
What I wanted to do next was to amplify the whole virus sequence and I thought it would be possible to do it using the 2 host primers I have. However, my boss told me that it was impossible but he asked me to figure out why by myself.
I asked some of my colleagues and our post-doc and we seem to think that it was theoretically possible to do that even though it will be challenging.
The thing is, my boss is stressing that it is impossible. The only hint he gave is that human have 46 chromosomes.
Can anyone help me to answer this question?
** I attach a picture below to make it easier to understand. My question is why can't I amplify the whole provirus with primers HF and HR. Theoretically, PCR preferentially amplifies shorter fragments, so the chromosome without the virus would be preferentially amplified. However, there is still a chance that the longer, larger fragment is also amplified, right? Furthermore, if I perform some sort of enrichment for the virus, using probes for example to pull down the segment containing the virus only then perform PCR, theoretically I will be able to amplify the virus fragment, am I right?
