February 10, 2011 at 1:27 am #14491
I just register as a new member.
By the way, I am a scientist in research department of an Industry
I will be working with genes, so now they put me in some kind of self-training on PCR etc
I will be working particularly with CYPs, and they gave me some literature on it.
I have read some other (literature) myself also, and I found that genes coding CYPs size in kbp. Surprsingly, all the project done in this lab concerning CYPs take 200-300 bp as CYP gene size. When I ask the other scientist worked on this project, she said that the size was taken from the primer designing.
We used NCBI help on designing primer. The final result should reveal your primer sequence and other information such as tm, %GC, etc including Product Size. This is where they took the size they’ve been working with.
And how about the size I found in literature? Which one should I use? Or they simply just a different term? But, still, which one should I use?
And which term indicates the possibility of your primer to form dimers?
Thanks a lot guys,
February 10, 2011 at 6:26 am #103472
200-300 bp would mean just 100 amino acids! That’s definitely not enough. Just two examples from my work:
http://arabidopsis-p450.biotec.uiuc.edu … 35A1.shtml
http://arabidopsis-p450.biotec.uiuc.edu … 35A2.shtml
you see, they have both several kbp.
However, question is, what you need it for. What they use these fragments in your job for.
February 10, 2011 at 7:47 am #103474
Thank your very much JackBean
Well, now i know that it is too short too be transcripted any further, 🙂
But what about that term "product size" written in primer result form? What do you think it refer to? Is it referring to primer size? But they write the sequence, and its only 21 bp long.
We want to check the effect of extract on CYP expression.
February 10, 2011 at 8:29 am #103475
I see. And you’re using real-time PCR, rigth?
February 10, 2011 at 8:57 am #103476
no, just reverse transcription and usual PCR.
As quantification, we will compare the expression to control (non treated cell)
February 10, 2011 at 9:14 am #103478
I see. Well, in that case, you don’t need whole-length gene, but only small fragment with primers to most diverse parts are the best.
February 10, 2011 at 9:18 am #103479
excuse me JackBean, but I dont understand your last sentence: …only small fragment with primers to most diverse parts are the best.
You mean the primers could anneal at many site and produce various sequence?
Thank you soo much
February 21, 2011 at 9:24 am #103574
sure, you can design primers to any part of your gene (or even outside of any gene :lol:)
- You must be logged in to reply to this topic.