Quantitation and Database

Transcriptomic arrays focus on the mRNA binding to its DNA transcription template, identifying the correct originating sequence to characterize gene expression. The signal intensity of the labeled mRNA bound correlates with the level of gene expression. However, a main disadvantage is that the length of the probe sequence and of the DNA target has to be long enough to eliminate non-specificity of binding, which nullifies the goal of the experiment since it relies on binding specificity.

A solution is to analyze the target sequence beforehand against known DNA sequence patterns to ensure that non-specificity of binding will be insignificant. Another method for mRNA quantitation is Real-Time PCR. Real-Time PCR aims at quantitating changes in gene expression. Fluorescent probes are used to visualize the gene expression levels. The design of the cyclic phase must be set so that data collection takes place during the amplification process to obtain a good strong intensity signal. At a specific cycle number, the reactions are analyzed by computer.

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The most common result analysis involves a standard curve that is used as a reference standard for extrapolating the quantitative information for the mRNA targets. Actually, RT-PCR is also used along with microarrays technology but can stand on its own as well using wellplates. (Boutros, 2004) diagram taken from TaqMan Gold RT-PCR kit protocol (PE Applied Biosystems) Taken from the Nucleic Acids Research Facilities, Virginia Commonwealth University http://www. narf. vcu. edu/pcr3. html The National Center for Biotechnology Information (NCBI) is the most expansive and used database in the U.

S. It was sponsored by Senator Claude Pepper who realized that computerized stored information was essential for progress in science. On November 14 1988, NCBI was born as a division of the National Library of Medicine (NLM) and the National Institute of Health (NIH). The advantage is that NLM had experience creating and maintaining medical databases while the NIH was used to create computational methods to store, retrieve and use the data. Data are: literature/Entrez/nucleotide genome-specific.

Software tools work for data mining, display 3D structures (PDB –Protein Data Bank among others), maps, pages for collaborations with cancer research, FTP downloading sites, and statistics. The bioinformatics components are submitted nucleotides or protein sequences by laboratories and data exchange with the European Molecular Biology Lab, and with the DNA database of Japan. http://www. ncbi. nih. gov/About/primer/index. html Reference Boutros PC, Okey AB. (2004) PUNS: transcriptomic- and genomic-insilico PCR for enhanced primer design. Bioinformatics 20 (15): 2399-2400.