d7880d4570194bd583252958b3bedce55184ed58 jnavarr5 Mon Apr 15 09:53:50 2019 -0700 Updating http to http for mm10, uiLinks cronjob. diff --git src/hg/makeDb/trackDb/qPcrPrimers.html src/hg/makeDb/trackDb/qPcrPrimers.html index 3a419b6..12c15e0 100644 --- src/hg/makeDb/trackDb/qPcrPrimers.html +++ src/hg/makeDb/trackDb/qPcrPrimers.html @@ -1,102 +1,102 @@ <H2>Description</H2> <P> This track provides automatically-designed RT-qPCR primers for measuring the abundance of human and mouse transcripts using SYBR-based qPCR (qPCR with double-stranded DNA-binding reporter dye). The primers were generated by a procedure that targets all transcripts and all "possible" exon-exon and intron-exon junctions in the human and mouse transcriptomes. </P> <P> Not all consecutive exon-intron-exon triplets generate "possible" primer pairs. "Possible" primer pairs are defined as satisfying a set of imposed design rules: <OL> <LI> The first exon-exon junction is not addressed, to avoid problems related to abortive transcription.</LI> <LI>Intron length should be more than 800 bp to avoid problems of double products in amplification.</LI> <LI>Only junction primers are designed: forward and reverse primers must flank the junction.</LI> <LI>Melting temperature of the primers should be between 60°C and 63°C (optimally 60.5°C, according to Breslauer <em>et al.</em>, 1986).</LI> <LI>Primer length should be 18-25 bp.</LI> <LI>Product size should be 60-125 bp.</LI> <LI>Primers are designed first for the intron-exon (pre-mRNA) junctions, and the two best primer pairs for each junction are chosen. <LI>Then, for the reverse primer of each pair, two options are designed for the corresponding forward primer of the exon-exon (mRNA) junction.</LI> <LI>The "Primer Mispriming Library" of the <A HREF="http://bioinfo.ut.ee/primer3-0.4.0/" TARGET=_blank>primer3</A> software is used: "human" for the human transcriptome and "rodent" for the mouse transcriptome.</LI> </OL> </P> <P> The track provides easy access to primers for almost all transcripts in the transcriptome, eliminating the need for a tedious, error-prone design process. </P> <H2>Methods</H2> <p> The <a href="../cgi-bin/hgGene?hgg_do_kgMethod=1&org=$organism">UCSC Genes model</a> was used as a reference of the gene structure and the <A HREF="http://bioinfo.ut.ee/primer3-0.4.0/" TARGET=_blank>primer3</A> software as the design engine. The software goes over all possible exon-exon junctions in the transcriptome and applies our design rules/parameters to provide two primer pairs for every "possible" intron-exon junction and four pairs for every "possible" exon-exon junction. </P> <H2>Display Conventions</H2> <P> The primers to amplify pre-mRNA (intron-exon junctions) are shown in <font color="red"><b>red</b> </font> and the primers to amplify mRNA (exon-exon junctions) in <font color="blue"><b>blue</b></font>. For each pre-mRNA primer pair, there are two corresponding mRNA primers (that use the same reverse primer, if possible). Each pair has a unique code which stands for the gene name and the junction name. For example, the human pair "JAG1_uc002wnw.2_11_1" amplifies pre-mRNA, and the corresponding mRNA primers are "JAG1_uc002wnw.2_11_1_1" and "JAG1_uc002wnw.2_11_1_2."</p> <p> Using <b>JAG1_uc002wnw.2_11_1_2</b> to illustrate the naming scheme: <ul> <li><b>JAG1</b> is the gene symbol. <li><b>uc002wnw.2</b> is the UCSC Genes identifier of the isoform. <li><b>11</b> identifies the exon-intron-exon triplet. <li><b>1</b> is the number (1 or 2) of the intron-exon junction (step 7 in the <em>Description</em> section above). The names of primer pairs that cover intron-exon junctions end here. For the exon-exon junctions that use the same reverse primer, there is one additional number. <li><b>2</b> is the number (1 or 2) of the exon-exon junction pair (step 8 in the <em>Description</em> section above). </ul> <p> Clicking on a primer pair will take you to a new page with details for that pair. Additional properties for the primer pair, including forward and reverse sequence, melting temperature, GC%, and product size, are available by clicking on the number next to the instruction "Click here for primer details." There is also a <A HREF="http://www.weizmann.ac.il/complex/compphys/software/Amit/primers/batch_query_qpcr_primers.htm" TARGET=_blank>batch query website</A> available to download details for a large number of primers. </p> <H2>References</H2> <p> Breslauer KJ, Frank R, Blöcker H, Marky LA. -<a href="http://www.pnas.org/content/83/11/3746" target="_blank"> +<a href="https://www.pnas.org/content/83/11/3746" target="_blank"> Predicting DNA duplex stability from the base sequence</a>. <em>Proc Natl Acad Sci U S A</em>. 1986 Jun;83(11):3746-50. PMID: <a href="https://www.ncbi.nlm.nih.gov/pubmed/3459152" target="_blank">3459152</a>; PMC: <a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC323600/" target="_blank">PMC323600</a> </p> <p> Rozen S, Skaletsky H. <a href="https://www.ncbi.nlm.nih.gov/pubmed/10547847" target="_blank"> Primer3 on the WWW for general users and for biologist programmers</a>. <em>Methods Mol Biol</em>. 2000;132:365-86. PMID: <a href="https://www.ncbi.nlm.nih.gov/pubmed/10547847" target="_blank">10547847</a> </p> <p> Zeisel A, Yitzhaky A, Bossel Ben-Moshe N, Domany E. <a href="https://academic.oup.com/bioinformatics/article/29/10/1355/260020/An-accessible-database- for-mouse-and-human-whole" target="_blank"> An accessible database for mouse and human whole transcriptome qPCR primers</a>. <em>Bioinformatics</em>. 2013 May 15;29(10):1355-6. PMID: <a href="https://www.ncbi.nlm.nih.gov/pubmed/23539303" target="_blank">23539303</a> </p>