One of my first posts on this blog was titled "Vector Difficulties". That was on the 3rd of March, 2005. It is now the 4rth of May, 2006 and it has taken me THIS LONG to get POSITIVE CLONES! I cannot beleive that I have spent 1 year, 3 months and 2 days on this!!!!
However, I think I have encountered EVERY SINGLE problem along the way! For the first few months, I was perfecting my techniques. Then towards the end of last year, I realised that I was actually using the wrong enzyme!!! So I started using the correct enzyme, and again NOTHING!
I decided to start from scratch again this year. This time I realised that there was a restriction site which I had completely forgotten about! Then my competent cells weren't competent, and when they were, I kept loosing my inserts and vectors!
Finally, I got it right!!! TODAY!
This morning:
SW: Have you looked at your paltes yet?
Me: No... it is just going to make me depressed, so I will wait till mid-day or so...
SW: Let me go and look for you!!
DC: She has been having some good cloning vibes lately you know!
Me: I know... but that doesn't mean anything for me!
I then walked over to check out my plates.
SW: Ok.... which plates are you expecting to see colonies on? (She was holiding them and I couldn't see...)
Me: Oh God! There is nothing again? Just one colony would be good on any plate other than the negative control!
SW: Hmmm
Me: Why won't cloning work for me??
SW: Well, there are colonies on your vector only...
Me: Huh?
SW: Hmm... I see more colonies...
Me: WHAT?
She then showed me my BEAUTIFUL plates and I squealed (a bit like I do when I sit in the front seat of a car and the driver goes around a round-a-bout REALLY FAST!)!! I was sooo excited to just see colonies!
I now know that I even have POSITIVE colonies! When I found that out, I was bursting with excitement and I jumped around for a bit and tired myself out... sad but true... :P
I am just going to enjoy this moment while it lasts! I still have to sequence verify and see if my construct will actually express, but YIIPPPEEEE!!!!! (For now...)
("Any old monkey can clone", was one of the first things someone said to me when I told them I was planning to clone...)
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