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Biotech / Medical : VD's Model Portfolio & Discussion Thread -- Ignore unavailable to you. Want to Upgrade?

To: Rocketman who wrote (5131)	6/17/1998 6:34:00 PM
From: poodle	Read Replies (1) \| Respond to of 9719

<<Any hybridization experts out there, we'd love to hear from you.>> Not sure you either need hybridization expert or would love to hear what s/he may say. :-) <<It's been 15 years since I personally hybridized DNA, but the longer the probe, the more specific it can be. Basically this comes down to the odds of having a specific 20 bp sequence being repeated in many genes, versus the odds of a 1000 bp sequence being repeated. As far as the better way to find SNPs, I'm not at all sure. INCY finds a lot of SNPs just in the sequencing itself. My gut feeling guess is that the longer the probe the better for finding SNPs, but I'm not totally sure on the point.>> I'm not totally sure, but may be you mixed up different aims, applications and technologies. "Longer the probe, the more specific it can be." Agree completely. That why long probes are used to determine an existence of specific molecule (DNA or RNA) in the sample. So, if you need to know what RNAs, for example, are expressed in tissue or cell type, you should hybridize cDNA from this tissue to the library of known sequences. If you have positive signal, gene expressed, if not, it's not. This way you will know which genes are expressed in any tissue you are interested in. Vice versa, after you hybridize your chip or filter to different cDNAs pools from different tissues you may know in which tissues (and at what stage) gene of interest is expressed. That's gene expression analysis. Short fragments are inappropriate for this type of analysis because of the specificity problem you mentioned. In case if you are searching for the mutations (let say differences between "normal" gene and gene from patient, for example) you can't use big pieces because they will hybridize to each other even if they are not completely identical. If you have 1000 nucleotides piece with 1 missmatch (0.1%), I seriously doubt you can find any simple hybridization technique to determine this difference. There are some ways to do that but they are more complex. If you have 20 nucleotides fragment and 1 mutation (5%) you definitely can find conditions that will allow hybridization of completely identical fragments, but not fragments with 1 different position. That's sequence polymorphism analysis. DNA for these two methods are created by two absolutely different tech. For the first one it will be PCR of cDNAs. For the second one oligonucleotide synthesizer should be used. I am not an expert here, have no any position or interest, just happened to run near by.