Guidance for array-based CGH - (Sep/13/2008 )
Hi...
I am new to microarray field but going to perform array-based CGH for my project.
I've been searching the informations from the internet since last week, and get to know bits of bits for this method. However, there are 2 things i am still blur...
First of all, i need to prepared a glass slide spotted with the PCR products of the genes of interest. In this case, there are ~4200 annotated genes for my reference bacterial, and i need to do PCR for ~4200 times to get the product???? and ~8400 oligonucleotide to order for PCR???
Next, I will need to prepare and label my target DNA. I will prepare gDNA for this purpose... is it ok?
Hope someone will help me to make it clear....
Thanks.
I dont think that they perform PCR. Normally you have only 70mers on the slides, not the full gene. And they are synthesized in the way how how you would make primers primers. However, there are companies who perform that task for you. And if you are lucky, the TIGR institute (now J. Craig Venter institute) has slides with your organism. That means you can get them for free (www.jcvi.org)
I am new to microarray field but going to perform array-based CGH for my project.
I've been searching the informations from the internet since last week, and get to know bits of bits for this method. However, there are 2 things i am still blur...
First of all, i need to prepared a glass slide spotted with the PCR products of the genes of interest. In this case, there are ~4200 annotated genes for my reference bacterial, and i need to do PCR for ~4200 times to get the product???? and ~8400 oligonucleotide to order for PCR???
Next, I will need to prepare and label my target DNA. I will prepare gDNA for this purpose... is it ok?
Hope someone will help me to make it clear....
Thanks.
Hello,
With a such project, you may find a prespotted microarray containg all your genes and cheaper than your planed costs ! Check your account and compare with market prices ! You would be suprised !
Sedondelly, it depends on your microarry type and purpose (gene expression profiling, genotyping, polymorphism...etc). For gene expression profiling microarray, I would say that you can amplify from genomic DNA if your PCR products were designed to be in 3' ou 5' regions and/or in exons regions. otherwise, you cant detect any expressed gene if your PCR product was designed in an intron region.
But, I thing it would be a tedisous task to amplify manually 4200 genes whereas there a many microarry facilities plateformes.
In your place, I'll check the prices and chose consequently.
Good luck
I am new to microarray field but going to perform array-based CGH for my project.
I've been searching the informations from the internet since last week, and get to know bits of bits for this method. However, there are 2 things i am still blur...
First of all, i need to prepared a glass slide spotted with the PCR products of the genes of interest. In this case, there are ~4200 annotated genes for my reference bacterial, and i need to do PCR for ~4200 times to get the product???? and ~8400 oligonucleotide to order for PCR???
Next, I will need to prepare and label my target DNA. I will prepare gDNA for this purpose... is it ok?
Hope someone will help me to make it clear....
Thanks.
Hello,
With a such project, you may find a prespotted microarray containg all your genes and cheaper than your planed costs ! Check your account and compare with market prices ! You would be suprised !
Sedondelly, it depends on your microarry type and purpose (gene expression profiling, genotyping, polymorphism...etc). For gene expression profiling microarray, I would say that you can amplify from genomic DNA if your PCR products were designed to be in 3' ou 5' regions and/or in exons regions. otherwise, you cant detect any expressed gene if your PCR product was designed in an intron region.
But, I thing it would be a tedisous task to amplify manually 4200 genes whereas there a many microarry facilities plateformes.
In your place, I'll check the prices and chose consequently.
Good luck
You should buy your arrays already printed. For aCGH we use in the lab, Agilent, Nimblegen and Affymetrix. It is cheaper and better quality.