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RT-PCR

experiment: RT-PCR project: The Drosophila Transcriptome PI: Sue Celniker Labs: Michael Brent
These experiments aim to annotate the genome of Drosophila melanogaster. PCR targets are selected based on existing evidence for predicted gene models. The latest ESTs, mRNAs and experimental data (if not already in Genbank) are collapsed into clusters, aligned, processed into multi-exon predicted transcripts, then classified into 3 main categories: known, novel, and partially novel. These categories are defined as follows: 1. Known: All introns are overlapped by EST or mRNA sequence 2. Partial: Some, but not all, introns are overlapped by EST or mRNA sequence 3. Novel: No introns are overlapped by EST or mRNA sequence These categories may be further sub-categorized depending on whether the predicted intron/exon structure is supported, whether the overlapping ESTs are spliced, etc. PCR primers are designed to amplify regions of partially and totally novel predictions which have unverified introns. In some cases, protein homology with other species is used to prioritize novel predictions for testing. An initial test set of completely novel predictions may have 35-75% homology to proteins of other Drosophila species, for example. PCR products are amplified by reverse transcription using Superscript III (Invitrogen) followed by standard PCR with Phusion polymerase, from pools of RNA from Cherbas et al. Products were cycle sequenced using both F and R pcr primers performed either in-house by the standard ABI Big Dye version 3.1 protocol or outsourced. The resulting EST is aligned to the dm3/Release 5 Drosophila genome using GMAP. An EST is considered a hit if it results in a spliced alignment with over 90 percent identity that aligns to the same locus as it was designed for, or if it aligns to a locus in the genome that is not covered by any transcript in the experiment. Positive EST sequences are submitted to genbank.

All features generated by this experiment:
Feature Export
CDNAMatch ? 2352
This represents 2349 unique CDNAMatchs
TAB CSV GFF3 (for UCSC) SEQ
MRNA ? 1691
This represents 15 unique MRNAs
TAB CSV GFF3 SEQ
EST ? 2352 TAB CSV GFF3 SEQ
All GBrowse tracks generated for this experiment:
GBrowse tracks: by chromosome
All entries in public repositories generated by this experiment
There are 5 RTPCR submissions for this experiment:
DCC id Name cell line Embargoed until Features, GBrowse and Data Files
modENCODE_214

DMG1 RTPCR embargo expired
? CDNAMatch: 572   export: TAB   CSV  GFF3  (for UCSC)  SEQ    create LIST
? MRNA: 431   export: TAB   CSV GFF3 SEQ    create LIST
? EST: 572   export: TAB   CSV GFF3 SEQ    create LIST
  [find nearby features]
GBrowse tracks: DMG1_RTPCR_cDNA_match_1278_214
DMG2_RTPCR_mRNA_1404_214
Trace Archive: 1967618070
Trace Archive: 1967618071
Trace Archive: 1967618072
Trace Archive: 1967618073
Trace Archive: 1967618074
...All entries in public repositories generated by this submission



1 files Download data files
modENCODE_339

DMG3 RTPCR embargo expired
? CDNAMatch: 322   export: TAB   CSV  GFF3  (for UCSC)  SEQ    create LIST
? MRNA: 224   export: TAB   CSV GFF3 SEQ    create LIST
? EST: 322   export: TAB   CSV GFF3 SEQ    create LIST
  [find nearby features]
GBrowse tracks: DMG3_RTPCR_mRNA_1429_339
DMG3_RTPCR_cDNA_match_1431_339
Trace Archive: 2086863590
Trace Archive: 2086863593
Trace Archive: 2086863594
Trace Archive: 2086863595
Trace Archive: 2086863596
...All entries in public repositories generated by this submission



1 files Download data files
modENCODE_340

DMG4 RTPCR embargo expired
? CDNAMatch: 536   export: TAB   CSV  GFF3  (for UCSC)  SEQ    create LIST
? MRNA: 312   export: TAB   CSV GFF3 SEQ    create LIST
? EST: 536   export: TAB   CSV GFF3 SEQ    create LIST
  [find nearby features]
GBrowse tracks: DMG4_RTPCR_mRNA_1452_340
DMG4_RTPCR_cDNA_match_1456_340
Trace Archive: 2179574459
Trace Archive: 2179574460
Trace Archive: 2179574461
Trace Archive: 2179574462
Trace Archive: 2179574463
...All entries in public repositories generated by this submission



1 files Download data files
modENCODE_215

DMG2 RTPCR embargo expired
? CDNAMatch: 508   export: TAB   CSV  GFF3  (for UCSC)  SEQ    create LIST
? MRNA: 440   export: TAB   CSV GFF3 SEQ    create LIST
? EST: 508   export: TAB   CSV GFF3 SEQ    create LIST
  [find nearby features]
GBrowse tracks: DMG2_RTPCR_cDNA_match_1412_215
DMG1_RTPCR_mRNA_1273_215
Trace Archive: 1968181244
Trace Archive: 1968181245
Trace Archive: 1968181246
Trace Archive: 1968181247
Trace Archive: 1968181248
...All entries in public repositories generated by this submission



1 files Download data files
modENCODE_2270


Related to: modENCODE_555

MB6 RTPCR CME L1 [ALL]
embargo expired
? CDNAMatch: 414   export: TAB   CSV  GFF3  (for UCSC)  SEQ    create LIST
? MRNA: 284   export: TAB   CSV  GFF3  (for UCSC)  SEQ    create LIST
? EST: 414   export: TAB   CSV GFF3 SEQ    create LIST
  [find nearby features]
GBrowse tracks: MB6_RTPCR_cDNA_match_642_2270
Trace Archive: 2239708162
Trace Archive: 2239708163
Trace Archive: 2239708164
Trace Archive: 2239708165
Trace Archive: 2239708166
...All entries in public repositories generated by this submission



1 files Download data files