intronEst Spliced ESTs C. briggsae ESTs That Have Been Spliced mRNA and EST Description This track shows alignments between C. briggsae expressed sequence tags (ESTs) in GenBank and the genome that show signs of splicing when aligned against the genome. ESTs are single-read sequences, typically about 500 bases in length, that usually represent fragments of transcribed genes. To be considered spliced, an EST must show evidence of at least one canonical intron, i.e. one that is at least 32 bases in length and has GT/AG ends. By requiring splicing, the level of contamination in the EST databases is drastically reduced at the expense of eliminating many genuine 3' ESTs. For a display of all ESTs (including unspliced), see the C. briggsae EST track. Display Conventions and Configuration This track follows the display conventions for PSL alignment tracks. In dense display mode, darker shading indicates a larger number of aligned ESTs. The strand information (+/-) indicates the direction of the match between the EST and the matching genomic sequence. It bears no relationship to the direction of transcription of the RNA with which it might be associated. The description page for this track has a filter that can be used to change the display mode, alter the color, and include/exclude a subset of items within the track. This may be helpful when many items are shown in the track display, especially when only some are relevant to the current task. To use the filter: Type a term in one or more of the text boxes to filter the EST display. For example, to apply the filter to all ESTs expressed in a specific organ, type the name of the organ in the tissue box. To view the list of valid terms for each text box, consult the table in the Table Browser that corresponds to the factor on which you wish to filter. For example, the "tissue" table contains all the types of tissues that can be entered into the tissue text box. Wildcards may also be used in the filter. If filtering on more than one value, choose the desired combination logic. If "and" is selected, only ESTs that match all filter criteria will be highlighted. If "or" is selected, ESTs that match any one of the filter criteria will be highlighted. Choose the color or display characteristic that should be used to highlight or include/exclude the filtered items. If "exclude" is chosen, the browser will not display ESTs that match the filter criteria. If "include" is selected, the browser will display only those ESTs that match the filter criteria. This track may also be configured to display base labeling, a feature that allows the user to display all bases in the aligning sequence or only those that differ from the genomic sequence. For more information about this option, go to the Base Coloring for Alignment Tracks page. Methods To make an EST, RNA is isolated from cells and reverse transcribed into cDNA. Typically, the cDNA is cloned into a plasmid vector and a read is taken from the 5' and/or 3' primer. For most — but not all — ESTs, the reverse transcription is primed by an oligo-dT, which hybridizes with the poly-A tail of mature mRNA. The reverse transcriptase may or may not make it to the 5' end of the mRNA, which may or may not be degraded. In general, the 3' ESTs mark the end of transcription reasonably well, but the 5' ESTs may end at any point within the transcript. Some of the newer cap-selected libraries cover transcription start reasonably well. Before the cap-selection techniques emerged, some projects used random rather than poly-A priming in an attempt to retrieve sequence distant from the 3' end. These projects were successful at this, but as a side effect also deposited sequences from unprocessed mRNA and perhaps even genomic sequences into the EST databases. Even outside of the random-primed projects, there is a degree of non-mRNA contamination. Because of this, a single unspliced EST should be viewed with considerable skepticism. To generate this track, C. briggsae ESTs from GenBank were aligned against the genome using blat. Note that the maximum intron length allowed by blat is 750,000 bases, which may eliminate some ESTs with very long introns that might otherwise align. When a single EST aligned in multiple places, the alignment having the highest base identity was identified. Only alignments having a base identity level within 0.5% of the best and at least 96% base identity with the genomic sequence are displayed in this track. Credits This track was produced at UCSC from EST sequence data submitted to the international public sequence databases by scientists worldwide. References Benson DA, Karsch-Mizrachi I, Lipman DJ, Ostell J, Wheeler DL. GenBank: update. Nucleic Acids Res. 2004 Jan 1;32(Database issue):D23-6. PMID: 14681350; PMC: PMC308779 Kent WJ. BLAT - the BLAST-like alignment tool. Genome Res. 2002 Apr;12(4):656-64. PMID: 11932250; PMC: PMC187518 
gold Assembly Assembly from Fragments Mapping and Sequencing Description This track shows the draft assembly of the C. briggsae genome. This version was assembled from 2.05 million whole genome shotgun (WGS) reads. The Phusion (Mullikin, J.) assembler was used to create contigs based on overlap information; supercontigs were then created using read pair information to cross gaps. These supercontigs were assembled into mapped ultracontigs based on FPC fingerprint mapping with material from previously finished clones used to bridge gaps. The sequence data was used to choose primers along major ultracontigs, from which a genetic map was constructed yielding six chromosomes. The genetic mapping data, along with data from 1:1 elegans:briggsae orthologs, were used to refine ultracontigs and to order and orient the sequences along the C. briggsae chromosomes. In dense mode, this track depicts the contigs that make up the currently viewed scaffold. Contig boundaries are distinguished by the use of alternating gold and brown coloration. Where gaps exist between contigs, spaces are shown between the gold and brown blocks. The relative order and orientation of the contigs within a scaffold is always known; therefore, a line is drawn in the graphical display to bridge the blocks. All components within this track are of fragment type "W": whole genome shotgun contig. 
augustusGene AUGUSTUS AUGUSTUS ab initio gene predictions v3.1 Genes and Gene Predictions Description This track shows ab initio predictions from the program AUGUSTUS (version 3.1). The predictions are based on the genome sequence alone. For more information on the different gene tracks, see our Genes FAQ. Methods Statistical signal models were built for splice sites, branch-point patterns, translation start sites, and the poly-A signal. Furthermore, models were built for the sequence content of protein-coding and non-coding regions as well as for the length distributions of different exon and intron types. Detailed descriptions of most of these different models can be found in Mario Stanke's dissertation. This track shows the most likely gene structure according to a Semi-Markov Conditional Random Field model. Alternative splicing transcripts were obtained with a sampling algorithm (--alternatives-from-sampling=true --sample=100 --minexonintronprob=0.2 --minmeanexonintronprob=0.5 --maxtracks=3 --temperature=2). The different models used by Augustus were trained on a number of different species-specific gene sets, which included 1000-2000 training gene structures. The --species option allows one to choose the species used for training the models. Different training species were used for the --species option when generating these predictions for different groups of assemblies. Assembly Group Training Species Fish zebrafish Birds chicken Human and all other vertebrates human Nematodes caenorhabditis Drosophila fly A. mellifera honeybee1 A. gambiae culex S. cerevisiae saccharomyces This table describes which training species was used for a particular group of assemblies. When available, the closest related training species was used. Credits Thanks to the Stanke lab for providing the AUGUSTUS program. The training for the chicken version was done by Stefanie König and the training for the human and zebrafish versions was done by Mario Stanke. References Stanke M, Diekhans M, Baertsch R, Haussler D. Using native and syntenically mapped cDNA alignments to improve de novo gene finding. Bioinformatics. 2008 Mar 1;24(5):637-44. PMID: 18218656 Stanke M, Waack S. Gene prediction with a hidden Markov model and a new intron submodel. Bioinformatics. 2003 Oct;19 Suppl 2:ii215-25. PMID: 14534192 
est C. briggsae ESTs C. briggsae ESTs Including Unspliced mRNA and EST Description This track shows alignments between C. briggsae expressed sequence tags (ESTs) in GenBank and the genome. ESTs are single-read sequences, typically about 500 bases in length, that usually represent fragments of transcribed genes. Display Conventions and Configuration This track follows the display conventions for PSL alignment tracks. In dense display mode, the items that are more darkly shaded indicate matches of better quality. The strand information (+/-) indicates the direction of the match between the EST and the matching genomic sequence. It bears no relationship to the direction of transcription of the RNA with which it might be associated. The description page for this track has a filter that can be used to change the display mode, alter the color, and include/exclude a subset of items within the track. This may be helpful when many items are shown in the track display, especially when only some are relevant to the current task. To use the filter: Type a term in one or more of the text boxes to filter the EST display. For example, to apply the filter to all ESTs expressed in a specific organ, type the name of the organ in the tissue box. To view the list of valid terms for each text box, consult the table in the Table Browser that corresponds to the factor on which you wish to filter. For example, the "tissue" table contains all the types of tissues that can be entered into the tissue text box. Wildcards may also be used in the filter. If filtering on more than one value, choose the desired combination logic. If "and" is selected, only ESTs that match all filter criteria will be highlighted. If "or" is selected, ESTs that match any one of the filter criteria will be highlighted. Choose the color or display characteristic that should be used to highlight or include/exclude the filtered items. If "exclude" is chosen, the browser will not display ESTs that match the filter criteria. If "include" is selected, the browser will display only those ESTs that match the filter criteria. This track may also be configured to display base labeling, a feature that allows the user to display all bases in the aligning sequence or only those that differ from the genomic sequence. For more information about this option, go to the Base Coloring for Alignment Tracks page. Methods To make an EST, RNA is isolated from cells and reverse transcribed into cDNA. Typically, the cDNA is cloned into a plasmid vector and a read is taken from the 5' and/or 3' primer. For most — but not all — ESTs, the reverse transcription is primed by an oligo-dT, which hybridizes with the poly-A tail of mature mRNA. The reverse transcriptase may or may not make it to the 5' end of the mRNA, which may or may not be degraded. In general, the 3' ESTs mark the end of transcription reasonably well, but the 5' ESTs may end at any point within the transcript. Some of the newer cap-selected libraries cover transcription start reasonably well. Before the cap-selection techniques emerged, some projects used random rather than poly-A priming in an attempt to retrieve sequence distant from the 3' end. These projects were successful at this, but as a side effect also deposited sequences from unprocessed mRNA and perhaps even genomic sequences into the EST databases. Even outside of the random-primed projects, there is a degree of non-mRNA contamination. Because of this, a single unspliced EST should be viewed with considerable skepticism. To generate this track, C. briggsae ESTs from GenBank were aligned against the genome using blat. Note that the maximum intron length allowed by blat is 750,000 bases, which may eliminate some ESTs with very long introns that might otherwise align. When a single EST aligned in multiple places, the alignment having the highest base identity was identified. Only alignments having a base identity level within 0.5% of the best and at least 96% base identity with the genomic sequence were kept. Credits This track was produced at UCSC from EST sequence data submitted to the international public sequence databases by scientists worldwide. References Benson DA, Karsch-Mizrachi I, Lipman DJ, Ostell J, Wheeler DL. GenBank: update. Nucleic Acids Res. 2004 Jan 1;32(Database issue):D23-6. PMID: 14681350; PMC: PMC308779 Kent WJ. BLAT - the BLAST-like alignment tool. Genome Res. 2002 Apr;12(4):656-64. PMID: 11932250; PMC: PMC187518 
mrna C. briggsae mRNAs C. briggsae mRNAs from GenBank mRNA and EST Description The mRNA track shows alignments between C. briggsae mRNAs in GenBank and the genome. Display Conventions and Configuration This track follows the display conventions for PSL alignment tracks. In dense display mode, the items that are more darkly shaded indicate matches of better quality. The description page for this track has a filter that can be used to change the display mode, alter the color, and include/exclude a subset of items within the track. This may be helpful when many items are shown in the track display, especially when only some are relevant to the current task. To use the filter: Type a term in one or more of the text boxes to filter the mRNA display. For example, to apply the filter to all mRNAs expressed in a specific organ, type the name of the organ in the tissue box. To view the list of valid terms for each text box, consult the table in the Table Browser that corresponds to the factor on which you wish to filter. For example, the "tissue" table contains all the types of tissues that can be entered into the tissue text box. Wildcards may also be used in the filter. If filtering on more than one value, choose the desired combination logic. If "and" is selected, only mRNAs that match all filter criteria will be highlighted. If "or" is selected, mRNAs that match any one of the filter criteria will be highlighted. Choose the color or display characteristic that should be used to highlight or include/exclude the filtered items. If "exclude" is chosen, the browser will not display mRNAs that match the filter criteria. If "include" is selected, the browser will display only those mRNAs that match the filter criteria. This track may also be configured to display codon coloring, a feature that allows the user to quickly compare mRNAs against the genomic sequence. For more information about this option, go to the Codon and Base Coloring for Alignment Tracks page. Methods GenBank C. briggsae mRNAs were aligned against the genome using the blat program. When a single mRNA aligned in multiple places, the alignment having the highest base identity was found. Only alignments having a base identity level within 0.5% of the best and at least 96% base identity with the genomic sequence were kept. Credits The mRNA track was produced at UCSC from mRNA sequence data submitted to the international public sequence databases by scientists worldwide. References Benson DA, Karsch-Mizrachi I, Lipman DJ, Ostell J, Wheeler DL. GenBank: update. Nucleic Acids Res. 2004 Jan 1;32(Database issue):D23-6. PMID: 14681350; PMC: PMC308779 Kent WJ. BLAT - the BLAST-like alignment tool. Genome Res. 2002 Apr;12(4):656-64. PMID: 11932250; PMC: PMC187518 
gap Gap Gap Locations Mapping and Sequencing Description This track depicts gaps in the assembly. These gaps - with the exception of intractable heterochromatic gaps - will be closed during the finishing process. Gaps are represented as black boxes in this track. If the relative order and orientation of the contigs on either side of the gap is known, it is a bridged gap and a white line is drawn through the black box representing the gap. This assembly contains the following principal types of gaps: Fragment - Gaps between the contigs of a draft clone. (In this context, a contig is a set of overlapping sequence reads.) Clone - Gaps between clones in the same map contig. 
gcPercent GC Percent Percentage GC in 20,000-Base Windows Mapping and Sequencing Description The GC percent track shows the percentage of G (guanine) and C (cytosine) bases in a 20,000 base window. Windows with high GC content are drawn more darkly than windows with low GC content. High GC content is typically associated with gene-rich areas. Credits This track was generated at UCSC. 
microsat Microsatellite Microsatellites - Di-nucleotide and Tri-nucleotide Repeats Variation and Repeats Description This track displays regions that are likely to be useful as microsatellite markers. These are sequences of at least 15 perfect di-nucleotide and tri-nucleotide repeats and tend to be highly polymorphic in the population. Methods The data shown in this track are a subset of the Simple Repeats track, selecting only those repeats of period 2 and 3, with 100% identity and no indels and with at least 15 copies of the repeat. The Simple Repeats track is created using the Tandem Repeats Finder. For more information about this program, see Benson (1999). Credits Tandem Repeats Finder was written by Gary Benson. References Benson G. Tandem repeats finder: a program to analyze DNA sequences. Nucleic Acids Res. 1999 Jan 15;27(2):573-80. PMID: 9862982; PMC: PMC148217 
xenoRefGene Other RefSeq Non-C. briggsae RefSeq Genes Genes and Gene Predictions Description This track shows known protein-coding and non-protein-coding genes for organisms other than C. briggsae, taken from the NCBI RNA reference sequences collection (RefSeq). The data underlying this track are updated weekly. Display Conventions and Configuration This track follows the display conventions for gene prediction tracks. The color shading indicates the level of review the RefSeq record has undergone: predicted (light), provisional (medium), reviewed (dark). The item labels and display colors of features within this track can be configured through the controls at the top of the track description page. Label: By default, items are labeled by gene name. Click the appropriate Label option to display the accession name instead of the gene name, show both the gene and accession names, or turn off the label completely. Codon coloring: This track contains an optional codon coloring feature that allows users to quickly validate and compare gene predictions. To display codon colors, select the genomic codons option from the Color track by codons pull-down menu. For more information about this feature, go to the Coloring Gene Predictions and Annotations by Codon page. Hide non-coding genes: By default, both the protein-coding and non-protein-coding genes are displayed. If you wish to see only the coding genes, click this box. Methods The RNAs were aligned against the C. briggsae genome using blat; those with an alignment of less than 15% were discarded. When a single RNA aligned in multiple places, the alignment having the highest base identity was identified. Only alignments having a base identity level within 0.5% of the best and at least 25% base identity with the genomic sequence were kept. Credits This track was produced at UCSC from RNA sequence data generated by scientists worldwide and curated by the NCBI RefSeq project. References Kent WJ. BLAT--the BLAST-like alignment tool. Genome Res. 2002 Apr;12(4):656-64. PMID: 11932250; PMC: PMC187518 Pruitt KD, Brown GR, Hiatt SM, Thibaud-Nissen F, Astashyn A, Ermolaeva O, Farrell CM, Hart J, Landrum MJ, McGarvey KM et al. RefSeq: an update on mammalian reference sequences. Nucleic Acids Res. 2014 Jan;42(Database issue):D756-63. PMID: 24259432; PMC: PMC3965018 Pruitt KD, Tatusova T, Maglott DR. NCBI Reference Sequence (RefSeq): a curated non-redundant sequence database of genomes, transcripts and proteins. Nucleic Acids Res. 2005 Jan 1;33(Database issue):D501-4. PMID: 15608248; PMC: PMC539979 
simpleRepeat Simple Repeats Simple Tandem Repeats by TRF Variation and Repeats Description This track displays simple tandem repeats (possibly imperfect repeats) located by Tandem Repeats Finder (TRF) which is specialized for this purpose. These repeats can occur within coding regions of genes and may be quite polymorphic. Repeat expansions are sometimes associated with specific diseases. Methods For more information about the TRF program, see Benson (1999). Credits TRF was written by Gary Benson. References Benson G. Tandem repeats finder: a program to analyze DNA sequences. Nucleic Acids Res. 1999 Jan 15;27(2):573-80. PMID: 9862982; PMC: PMC148217 
twinscan Twinscan Twinscan Gene Predictions Using C. briggsae/C. elegans WS98 Homology Genes and Gene Predictions Description The Twinscan program predicts genes in a manner similar to Genscan, except that Twinscan takes advantage of genome comparisons to improve gene prediction accuracy. More information and a web server can be found at http://mblab.wustl.edu/. Display Conventions and Configuration This track follows the display conventions for gene prediction tracks. The track description page offers the following filter and configuration options: Color track by codons: Select the genomic codons option to color and label each codon in a zoomed-in display to facilitate validation and comparison of gene predictions. Click the Help on codon coloring link on the track description page for more information about this feature. Methods The Twinscan algorithm is described in Korf, I. et al. (2001) in the References section below. Credits Thanks to Michael Brent's Computational Genomics Group at Washington University St. Louis for providing these data. References Korf I, Flicek P, Duan D, Brent MR. Integrating genomic homology into gene structure prediction. Bioinformatics. 2001;17 Suppl 1:S140-8. PMID: 11473003 
rmsk RepeatMasker Repeating Elements by RepeatMasker Variation and Repeats Description This track was created by using Arian Smit's RepeatMasker program which screens DNA sequences for interspersed repeats and low complexity DNA sequences. The program outputs a detailed annotation of the repeats that are present in the query sequence (represented by this track) as well as a modified version of the query sequence in which all the annotated repeats have been masked (generally available on the Downloads page). RepeatMasker uses the Repbase Update library of repeats from the Genetic Information Research Institute (GIRI). Repbase Update is described in Jurka, J. (2000) in the References section below. Display Conventions and Configuration In full display mode, this track displays up to ten different classes of repeats: Short interspersed nuclear elements (SINE), which include ALUs Long interspersed nuclear elements (LINE) Long terminal repeat elements (LTR) which include retroposons DNA repeat elements (DNA) Simple repeats (micro-satellites) Low complexity repeats Satellite repeats RNA repeats (including RNA, tRNA, rRNA, snRNA, scRNA) Other repeats which includes class RC (Rolling Circle) Unknown The level of color shading in the graphical display reflects the amount of base mismatch, base deletion, and base insertion associated with a repeat element. The higher the combined number of these, the lighter the shading. Methods UCSC has used the most current versions of the RepeatMasker software and repeat libraries available to generate these data. Note that these versions may be newer than those that are publicly available on the Internet. Data are generated using the RepeatMasker -s flag. Additional flags may be used for certain organisms. Repeats are soft-masked. Alignments may extend through repeats, but are not permitted to initiate in them. See the FAQ for more information. Credits Thanks to Arian Smit, Robert Hubley and GIRI for providing the tools and repeat libraries used to generate this track. References Repbase Update is described in Jurka J. Repbase update: a database and an electronic journal of repetitive elements. Trends Genet. 2000 Sep;16(9):418-420. 
chainCe2 Elegans Chain C. elegans (Mar. 2004 (WS120/ce2)) Chained Alignments Comparative Genomics Description This track shows alignments of C. elegans (ce2, Mar. 2004 (WS120/ce2)) to the C. briggsae genome using a gap scoring system that allows longer gaps than traditional affine gap scoring systems. It can also tolerate gaps in both C. elegans and C. briggsae simultaneously. These "double-sided" gaps can be caused by local inversions and overlapping deletions in both species. The chain track displays boxes joined together by either single or double lines. The boxes represent aligning regions. Single lines indicate gaps that are largely due to a deletion in the C. elegans assembly or an insertion in the C. briggsae assembly. Double lines represent more complex gaps that involve substantial sequence in both species. This may result from inversions, overlapping deletions, an abundance of local mutation, or an unsequenced gap in one species. In cases where multiple chains align over a particular region of the C. briggsae genome, the chains with single-lined gaps are often due to processed pseudogenes, while chains with double-lined gaps are more often due to paralogs and unprocessed pseudogenes. In the "pack" and "full" display modes, the individual feature names indicate the chromosome, strand, and location (in thousands) of the match for each matching alignment. Display Conventions and Configuration By default, the chains to chromosome-based assemblies are colored based on which chromosome they map to in the aligning organism. To turn off the coloring, check the "off" button next to: Color track based on chromosome. To display only the chains of one chromosome in the aligning organism, enter the name of that chromosome (e.g. chr4) in box next to: Filter by chromosome. Methods Transposons that have been inserted since the C. elegans/C. briggsae split were removed from the assemblies. The abbreviated genomes were aligned with blastz, and the transposons were then added back in. The resulting alignments were converted into axt format using the lavToAxt program. The axt alignments were fed into axtChain, which organizes all alignments between a single C. elegans chromosome and a single C. briggsae chromosome into a group and creates a kd-tree out of the gapless subsections (blocks) of the alignments. A dynamic program was then run over the kd-trees to find the maximally scoring chains of these blocks. Chains scoring below a threshold were discarded; the remaining chains are displayed in this track. Credits Blastz was developed at Pennsylvania State University by Minmei Hou, Scott Schwartz, Zheng Zhang, and Webb Miller with advice from Ross Hardison. Lineage-specific repeats were identified by Arian Smit and his RepeatMasker program. The axtChain program was developed at the University of California at Santa Cruz by Jim Kent with advice from Webb Miller and David Haussler. The browser display and database storage of the chains were generated by Robert Baertsch and Jim Kent. References Chiaromonte F, Yap VB, Miller W. Scoring pairwise genomic sequence alignments. Pac Symp Biocomput. 2002:115-26. PMID: 11928468 Kent WJ, Baertsch R, Hinrichs A, Miller W, Haussler D. Evolution's cauldron: Duplication, deletion, and rearrangement in the mouse and human genomes. Proc Natl Acad Sci U S A. 2003 Sep 30;100(20):11484-9. PMID: 14500911; PMC: PMC208784 Schwartz S, Kent WJ, Smit A, Zhang Z, Baertsch R, Hardison R, Haussler D, Miller W. Human-Mouse Alignments with BLASTZ. Genome Res. 2003 Jan;13(1):103-7. PMID: 12529312; PMC: PMC430961 
blastzCe2 Elegans Blastz C. elegans (Mar. 2004 (WS120/ce2)) Blastz Comparative Genomics Description This track displays blastz alignments of the C. elegans assembly (ce2, Mar. 2004 (WS120/ce2)) to the C. briggsae genome. The track has an optional feature that color codes alignments to indicate the chromosomes from which they are derived in the aligning assembly. To activate the color feature, click the on button next to "Color track based on chromosome" on the track description page. Using the Filter This track has a filter that can be used to change the display mode, turn on the chromosome color track, or filter the display output by chromosome. The filter is located at the top of the track description page, which is accessed via the small button to the left of the track's graphical display or through the link on the track's control menu. Color track: To display the chromosome color track, click the on button next to "Color track based on chromosome". When the color track is activated, each of the items within the annotation track will be colored to show the chromosome in the aligning genome assembly from which the alignment originated. Chromosome filter: To display only alignments from a specific chromomsome in the aligning assembly, type the chromosome number (in the form chrN) in the text box to the right of "Filter by chromosome". For example, to display alignments from chromosome 6, type "chr6". When you have finished configuring the filter, click the Submit button. Credits These alignments were contributed by Scott Schwartz of the Penn State Bioinformatics Group. The best-in-genome filtering was done using UCSC's axtBest program. References Chiaromonte F, Yap VB, Miller W. Scoring pairwise genomic sequence alignments. Pac Symp Biocomput. 2002:115-26. PMID: 11928468 Schwartz S, Kent WJ, Smit A, Zhang Z, Baertsch R, Hardison R, Haussler D, Miller W. Human-mouse alignments with BLASTZ. Genome Res. 2003 Jan;13(1):103-7. PMID: 12529312; PMC: PMC430961