Merlin is on track

Work marches on slowly. I did a PCR of wild type and a mutant (serine to alanine at position 518) Merlin. I actually got a band on the gel at the correct place but was still not sure if I successfully added the correct restriction enzyme sites to the 5' and 3' ends. So, I sent the PCR products in for sequencing and lo and behold, 8 of the 10 sequences were correct. Cause for a minor happy dance. Next, I will do a DNA clean up reaction. This will essentially filter out all the extra stuff from the PCR reaction such as residual enzyme and salts, leaving me with pure DNA. I can then proceed to do a restriction digest of both my PCR products (to "rough up" the ends) and of the viral vector that I will be dropping the PCR products into.

I just hired a student to work for me for the summer. He has no lab experience but is working out well. He is smart and is a hard worker. He has already gotten some results this week on a project that my last student (who just graduated and is going on to dental school) started. It is a pretty simple project but has been giving us headaches. One of the limitations of our work is the availablity of tumor samples. Many labs get around that problem by using cell lines. Cell lines are great in that they can be propagated for years and years (HeLa cells are cervical cancer cells that have been around since the 1950s) and are available whenever they are needed. On the flip side, immortalized cell lines are also a screwed up. You have essentially taken what in most cases are normal cells and made them into cancer cells so that they will continue to grow. This means that your results need to presented with the caveat that what you are reporting may not necessarily be exactly what happens in vivo. We all accept that and continue on with our work. We would like to be able to work with a cell line because we wouldn't have to be so dependant on someone getting cancer. However, there are no good vestibular schwannoma cell lines. As such we decided to make our own. We have two cell lines that we are working with and as part of the characterization process we are attempting to sequence the gene that encodes Merlin. We have isolated genomic DNA and are in the process of PCRing managable parts of the gene for sequencing. We have been having problems finding the right conditions but yesterday we actually got it right. With one part down, we have about 10 more to go. Let's keep our fingers crossed.