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Help -designing primers for real time RT-PCR - (Jun/04/2008 )

Hi all,

I want to design a primer pair for PORCINE (pig) model to analyze the gene expression of HSPA2 and several other genes.

1) when I go to nucleotide sequences of these genes it says mRNA linear, but do they correspond to cDNA?? is it that mRNA sequence always refer to cDNA? I am confused, please help with a explanation.

2) If I cant find cDNA sequence but 3 ESTs are available for HSPA2. can I use these sequences to design primers for Ral time RT-PCRs??


Ok I found out that all mRNA sequences in NCBI are actually cDNA sequences! blush.gif

can anyone give an answer to my second question??

The mRNA sequences stored in NCBI's sequence databases are really complementary DNA (cDNA) sequences generated from mRNA transcripts extracted from the cells of different organisms. Genomic DNA sequences of eukaryotic organisms contain exons (segments that encode proteins) interspersed with introns (noncoding segments or junk DNA). After transcription, introns are spliced out, exons are pieced together, and the mRNA strand is processed to form messenger RNA (mRNA). The removal of introns from mRNA sequences make cDNA much shorter than their genomic precursors.