Hidden ORF's in Trinity genes no reported by Transdecoder

[Cesar Hernandez]

Hi Brain 

I have a doubt whit some outputs in transdecoder program 

I have this trinity transcript

>TRINITY_DN177_c1_g1_i4 len=9235 path=[0:0-7518 3:7519-7786 6:7787-7812 7:7813-7825 9:7826-7828 11:7829-7849 12:7850-7893 14:7894-7901 18:7902-7948 20:7949-8286 22:8287-9133 31:9134-9165 33:9166-9234]

GAGATAATTATGGTCATATTATGAGATAATTATGGTCATATTATGAGATAATTATGGTCATATTATGCTACGAATCTGTGTCTATATTGGTGAATTTACCATGAAAAAGTGATATTTCCGGTACATGCCATTGAACGGCTTGGCTTACCTTCTCAATTATCGTGCTTGGTTTAAACGTTTCTTTTGTTCCGCTTCTATTTTGTTGTACTTTTCGCGCGAGGAACAAGGTTTTTTTCCTTTGCCTAAATATTTGCCTTTGGGTTTTGGTCCTCCAGAGAATATCACGTACTATGGCAGCGAAAGGAGCTTTAAGGTTTTAATTACCCCATAGCCATAGATTCTACTCGGTCTATCTATCATGTAACACTCCGTTGATGCGTACTAGAAAATGACAACGTACCGGGCTTGAGGGACATACAGAGACAATTACAGTAATCAAGAGTGTACCCAATTTTAACGAACTCAGTAAAAAATAAGGAATGTCGACATCTTAATTTTTTATATAAAGCGGTTTGGTATTGATTGTTTGAAGAATTTTCGGGTTGGTGTTTCTTTCTGATGCTACATAGAAGAACATCAAACAACTAAAAAAATATTATAATATGAAAAATATCATTTCATTGGTAAGCAAGAAGAAGGCTGCCTCAAAAAATGAGGATAAAAACATTTCTGAGTCTTCAAGAGATATTGTAAACCAACAGGAGGTTTTCAATACTGAAAATTTTGAAGAAGGGAAAAAGGATAGTGCCTTTGAGCTAGACCACTTAGAGTTCACCACCAATTCAGCCCAGTTAGGAGATTCTGACGAAGATAACGAGAATATGATTAATGAGATGAACGCTACTGATGAAGCAAATGAAGCTAACAGCGAGGAAAAAAGCATGACTTTAAAGCAGGCGTTGCTAAAATATCCAAAAGCAGCCCTGTGGTCCATATTAGTGTCTACTACCCTGGTTATGGAAGGTTATGATACCGCACTACTGAACGCACTGTATGCCCTGCCAGTGTTTCAGAGAAAATTCGGTACTTTGAACGGGGAGGGTTCTTACGAAATTACTTCCCAATGGCAGATTGGTTTAAACATGTGTGTCCAATGTGGTGAGATGATTGGTTTGCAAATCACGACTTATATGGTTGAATTTATGGGGAATCGTTATACGATGATTACAGCACTTGGTTTGTTAACTGCTTATATCTTTATCCTCTACTACTGTAAAAGTTTAGCTATGATTGCTGTGGGACAAGTTCTCTCAGCTATGCCATGGGGTTGTTTCCAGGGTTTGACTGTTACTTATGCTTCGGAAGTTTGCCCTTTAGCATTAAGATATTACATGACCAGTTACTCCAACATTTGTTGGTTATTTGGTCAAATCTTCGCCTCTGGTATTATGAAAAACTCACAAGAGAATTTAGGGAACTCTGACTTGGACTATAAATTGCCATTTGCTTTACAATGGATTTGGCCTGCTCCTTTAATGATCGGTATCTTTTTCGCTCCTGAGTCGCCCTGGTGGTTGGTGAGAAAGGATAGGGTCGCTGAGGCAAGAAAATCTTTAAGCAGAATTTTGAGTGGTAAAGGCGCCGAGAAGGACATTCAAGTTGATCTTACTTTAAAGCAGATTGAATTGACTATTGAAAAAGAAAGACTTTTAGCATCTAAATCAGGATCATTCTTTGATTGTTTCAAGGGAGTTAATGGAAGAAGAACGAGACTTGCATGTTTAGCTTGGGTAGCTCAAAATACTAGCGGTGCCTGTTTACTTGGTTACTCGACATATTTTTTTTGAAAGAGCAGGTATGGCCACCGACAAGGCGTTTACTTTTTCTGTAATTCAGTACTGTCTTGGGTTAGCGGGTACACTTTGCTCCTGGGTAATATCTGGCCGTGTTGGTAGATGGACAATACTGACCTATGGTCTTGCATTTCAAATGGTCTGCTTATTTGTTATTGGTGGAATGGGTTTTGGTTCTGGAAGCGGCGCTAGTAATGGTGCCGGTGGTTTATTGCTGGCTTTATCATTCTTTTACAATGCTGGTATCGGTGCAGTTGTTTACTGTATCGTAGCTGAAATTCCATCAGCGGAGTTGAGAACTAAGACTATAGTGCTGGCCCGTATTTGCTACAATCTCATGGCCGTTATCAACGCTATATTAACGCCCTATATGCTAAACGTGAGCGATTGGAACTGGGGTGCCAAAACTGGTCTATACTGGGGTGGTTTCACAGCAGTCACTTTAGCTTGGGTCATCATCGATCTGCCTGAGACAACTGGTAGAACCTTCAGTGAAATTAATGAACTTTTCAACCAAGGGGTTCCTGCCAGAAAATTTGCATCTACTGTGGTTGATCCATTCGGAAAGGGAAAAACTCAACATGATTCGCTAGCTGATGAGAGTATCAGTCAGTCCTCAAGCATAAAACAGCGAGAATTAAATGCAGCTGATAAATGTTAAGTAAAAGGGTTGTTTTTTTTTTTTGGAAGAAATAAGGAATCCCTTTGACTGCTCCCAAAACCCTCAGCTAGCTCGAGATTTTATATTTATACATTTTTTATTTTTCTGTAAAACATTTATATTTACCATTTTTTAAGCAAAATATTGTTAGTAGTTAGTTAAAATAGCCCAAGCAGCAATCAAGCAAATATGAGAGTATTTTTTCTTTAGCACCTGGTACTTGTGCCTGGATATTGATTCGAACAACATGCCAGGTCAACCGTATTCTCAATTAACTGTTACTTTAAATGTCCATACACTTAATAAAAAGAAGGAGAGAATCACGCAAAAATCACATTAAACTATATTATAATGTTTTATGAAGTATTTTTGAAGCATTAATGTGAGAGCTTATAGAGATACGGCATTCATAATTCAATGCTTTATTGGGATATACTATGCATATTCTATATGGTTCATGAAATATCTCAAAACATATCGCATAGTTTCACAATCGCTTGGCAAATAATTTTCCAATTCTGAGGATATAAACAACCATTTCAAAGAATTATTCTTTCTTGGTGACATGAAAATAATAATAAATTGCATTTGATATACAAAAGCGATTAGTAAAAAAAATCAAACCCATGTGGTCGTCATCCTCACAAACAATTAGGTAAATTTCGTGTTAGCAGTATGTGGTGAAGGTGTTCTAATTAGAAATTGTGATTCAAAAATCTATCACTTTGAATTTGAATATAATGGAATTGTTTTATAAATATTACCTAATTAGCCGCCACTGATATGGCTTCTTTGATCTATTTTTCAGTATAAAGTCAAACAAGATTTCAAAAAACCGGAGGAAGAATATGGGAAATCTACTAGTGTACAAGTTCGTCAAGTTTTCAATGGTCTGACAGCCCTCATCTGCCTTGTGAGAGGCTATAAAGTCGTTAGTCTTCATGTGGCGTATTACTTTTCTTCAGTAGATATTGTTATTATTACTAAGAATAGAGATATATATTGCATTATTGTGAAAAACATCGCTTCTTTTCTAATTTCCCCGCGTCAAGGGCTACGTGTTTTCTGTGAGATGTAAACCATATATCTAAAACAAGTATCATCACGCTTCTTTTATTTGTAGTATTTTTTCTTCGGTTTTCACTGGGGAGCCTGCCCAAGGCGAGGTGCAGTGTTCTTTCTACGGACTTTTTAGGAAGGAAATGAATTAAGCTACGCAGAAAGGACATCTCTTTTCAAAAGTTCTAGACATTTTCAGCCTTCATAGTTCCTAGAGATATGTCCGTATCTAGTAACTACTATAGTTAAATTTAGATCGGATTCTATTAATGTTATGTTCTGCCATAAAAATGTTGCTTTAAATGGCAAACATAAATGACAATAATATCTTGCCTCCTAAACTTAACTGGCATTTATCTATCTTCTCTGTATTGAATAAACGAATGGTGATTCACTCAAATATTCAAATCCCAGAGGTAGCAACATGGTAGCAGAAAGTACCGCCCCTTTCTTACCAATGCTGAATACACCATTTCTAAGTAATATTATCAACAAACCATAGCACTTATCTGCACTCAGCTGTTGTATTTCACTTGTTCCTCTACCCTTCAACTAGTGCTCCATTATAACGGTCGAGCAATACTTCATATCATTCATTTGAGCGTCAGTGCTGACGAAAGTATGATTTCATTAGCTACAATGTGGAAATATAATACTTCTCAAACACTGAAAAATTTCAAAACTTTATTATACAGTTCTTCAACAATAGTATGAGGTAATTACATGGCACTAAGTGCTTCAACGCGCTGTATATAAAGTCATCTTGATTATGCTAATTAAATCACAAAAGCGAAAGGATGAGAGTGAAAGTATCCATATTGCAGCTAGCCTATTCAACCTAAACTTGTCTCCCATCAGTACACAATCATCTCATCTAATATGGCTCTTAGCACTCAGTAAAAAAGTGTATATATACTGAAAACTGTTAGTATAGGAATTCTGGGAAGAATGCTAACGTTAGCGTGCCTAAAAGTATTTTATTAAATATAATTTTCAATAGGGAGACATCATGGTAAGAAGCAGTCTTATACTTGTAAAAGTAGACCTCCAAATAGAAAATTAATTACTCTGTTCATGAAAAGGGGTCGCAATTAGCAGTGAATTTTACCCTTGTGTTGCGGAAACACAGTTTCTACAATGGAAAGCTATCACTTACCAGGAAGTTGAGATGGACAAAAATGAAAATTAATCAAGGGTCTATGTCTTCATTATCCTTGGGATAACCATCCAATTGTAAAGGTTTAGAAATGGGCAGAGTAATTAGGGCATTCTGGCATTTGGTGGAAAATCTCTTCAGCATATCATTGTTATAGTGGTTTAAAGAAAAAGCAAATTTGCATACATCATGCAAAACATTCCATGCTTCTAATTTCATGTTATGATCATACTGGCCTACAATGTCCACCAAAGCAGTAGCTATTTCTAATGTTTTCACTGGTACCCCGGGACCATGGACATCGTAAAGATTCTCTGGGGTTAAGTAAGTATCTATTAACATATCTCTCGCAATTTCTATTGGTATTTGCCCATTGTTAGCGTTCAATGAAAAGTTGCTACCCCTCATTTGATAAGCTAGCTTCCATGCTAGTATCCTAATCCAACGCCGGGAAAATGAGATGTCTATGTAACCGTTAGACCATGGCTCTATTTCCGAGGAAGTTGTGTGGAGTTCGTTCCATATCTTTTTCAATGACTCTTCAGTGCAAGAAGCATCTGTAGAGTCAGCGGCTAAAGCATCGAAGAAACATTTTCCCGGCACGGTAAATATCCTAATCATTTCAAGGAAACTGTCCATAGAAAGCTGAGGATCAGTTACAAGTTCAAGTTGCGGTGGTGCTATTGTGGCATCCAGGCTCGTCGCACAATGAAGATATATGGCATAGTATCTCTCCGTCATGAGAAGCAAGTAATAAAGTTTCCGTCTAAGTTGCTGTTCTTCAAATGTAAGGGATCCGTAAGTTTCTTCCCGATGTAACCCTGCTACCGTAATCAGACCGACCGCTTCACAACAGAGTCTGTAAGAAGTTCTTGCGTTCGATATTTGTGCAAAGCTACGATGCAAACAGTAGTACGTCATAATATTGAATATATTGCTGTTATCCAAATCGTCGAATTGCTGACACGATGAGATGCAAAGATTAGATAACTGTTTTCCCGTGAAAGTGACTTCCTCTTCAGATTTTATTTCAGTTTGTAAATCACTGAGGGTGGCCGCTGATAAAGCGGTCAGAAACCAATATACGTAATTGTCATTGTATTATTCCTCCAGAAGTTTGTGAAGGTCATCGTACGAAAGAAGGGGCCATATTACGTATAAATTATCGTGATAGAGCCGCAAGCACTGATCGATTAGCTTTTTGGGAACCCTCTTATATATTACAGGAGCAGTTGCAATGGTGTTAGGACCGCTTTCCCTCTGCACTTCTGCTATTCTTTTCAAGCTCCTCAACCTAATGGACTTCGGACCTCTTTTTCTCGACGGTTGCAGATAAGTGCAATCCAAACTATTCTGTAGGCAACTGCTACACGGCCTTTTACCATCGCATTTCACTCGACGAATACGACAGCAGTCGCATGCCTGCTTGGCGCATGTTTGCTTAGTTAAAGTCATACTCGAAGCAAAAAAGAGTTCTGGCTAAATTTTCTACTTTTCATTTACTTTAATATATATCGAGATTATTAATTTTCTCTATCTGCGTACTTGAGTTATGAAGAATATAGCAAAATGAGTAATTATGACCGGGGATAAAGAAAACCCATTGTGCTAGCATATGGGGGCTGGTGGTGCAGGAAACCCAGGTCACCCGGACTTTATATGCCAACGTAACCTGCAGTCACCACTATTACTTGTTCAAGATTTTTATTTTGCTCTGCTTATTTCCATCTAAAATGAGAAACCTCAAGATCCGGTTGGAACGGGGCTGTATGTTTATGATTGCTCGAAGACCGTTATAATCACGTTTTTCTTGTTTTTTCGTCAAGAATTCCAGTCAAGTTTTCCATCACCTTGACCCTTAAAGCATCGACTTTTGTGCTCTTGAATGTGTTTCTAAGAATACTTGTAAAGGACACCCTCTAATTTCGTGTGCACTTTTCACATATTATCAAGACAATCGTTCCTGTACTCAGATGCACTGTTACTGTAAAGACTACTATACAACAAGCGAAAAATGATGTTCGAAAACCTTTATTTCTATTTTGAAAGGCATGTGTCTCGAGGTCCTTGCTTTATTGTGGGTGGTCATGCCATTCTGTAAACCTTACGGTACTGCTCCGTCTATATCTTTGAGGTTGTTATTTCCCCACAGATATGCGTTTCTAACCGAATATTCATTCAGTCGGACCGGACAATAGCTCTTAACTTCGTTTACCGGAGTAAATATCGTAAGAATTTGCATGCGGTGAAATACAGGGAAAATAAGAAATTACACCCTAATACAAAAAGAAAACTAAGTTTCACAATACGTAAGGATATTTTAGTGGGGAGAATATTTCGGAGAATAAAGTTTCCAACTCCGCGGTGTGAACAACCGCTCAGCACGCAGCGTTATTCTCGAGAAAAGTGGCCCTGAAATAAGGAGATAAAGTTACTAATGTTTTTTCGCTGTACGATATCAAATGTGACGAAGTAGGCACCCCACGCTATAAACTGGCTACTAAAGTTTATGTCAGTACTTGGGATCGTTGAAATACTCGGATAAATTATGTTCCTTATTTTTCATGGTTTTCGTCATACCACAGTTTACCCCAGAATGAGAAAGGATCTCCTTTTGAAATAAAAAGTACTTAAGGGCAATGATATTGAGTTGCTAGACGTTTGGTTAGACGCCTGTTTTGAAATAAAAAAGCTGTCTCAAATTAATCGAGCAAGCACAGATCAAACAAGATACAAACAAAGCTTTTCAACGTAATATTTACTATCGATGACTATTTCTTCTGCACATCCAGAGACAGAACCAAAGTGGTGGAAAGAGGCCACGTTCTATCAAATTTACCCAGCAAGTTTCAAAGACTCTAATGACGATGGCTGGGGTGATATGAAGGGTATTTCCTCCAAGTTGGAGTATATCAAGGAGCTTGGTGTCGATGCCATTTGGATCTCACCATTCTACGACTCGCCACAAGATGATATGGGTTACGATATTGCCAACTACGAAAAGGTCTGGCCAACCTACGGTACGAACGAGGACTGCTTTGCCTTGATCGAAAAGACACATAAGCTCGGTATGAAATTTATCACTGACTTGGTCATCAATCACTGTTCCAGCGAACATGAATGGTTCAAAGAGAGCAGATCCTCAAAAACCAATCCAAAACGTGACTGGTTCTTCTGGAGACCTCCTAAGGGTTATGACGCCGAAGGCAAGCCAATTCCTCCAAACAATTGGAGGTCTTACTTCGGTGGTTCTGCATGGACGTTTGATGAAAAGACACAAGAGTTTTACTTGCGTTTGTTTTGCTCCACCCAACCTGATCTCAACTGGGAAAACGAAGACTGCAGAAAGGCAATCTACGAAAGCGCTGTTGGATACTGGTTAGACCACGGTGTTGACGGTTTCAGAATCGATGTGGGGAGTTTGTACTCCAAAGTTGTTGGTCTACCAGATGCTCCTGTGATTGACAAGAACACAAAATGGCAGGCCAGTGATGCTTTCACAATGAATGGACCACGTATTCATGAATTCCACCAAGAAATGAACAAGTTCATGAGAGACAGGGTCAAAGATGGAAGAGAAATCATGACGGTTGGTGAAATGCAACATGCTTCCGACGAGACCAAGAAGTTGTACACGAGTGCCTCAAGACACGAGCTTAGTGAGCTGTTCAACTTTTCTCACACTGATGTTGGAACTTCTCCCAAGTTCCGCCAAAACTTGGTCCCATTTGAATTGAAGGATTGGAAAGTTGCTCTTGCCGAGCTGTTCAGTTATGTTAATGGAACTGATTGTTGGTCGACTATTTACCTAGAAAATCATGACCAACCTCGTTCGATTACGAGATTTGGTGACGACTCGCCCAAGAATCGTGTCGTTTCTGGTAAGTTGCTGTCTGTATTGTTAGTGTCACTGACCGGTACTTTGTATGTGTACCAGGGACAGGAGCTGGGCCAAATCAATTTCAAAAACTGGCCTATTGAAAAGTACGAGGACGTTGAAATCAGAAACAACTACAAGGCGATCAAGGAAGAGCATGGGGAAAACTCGAGCGAGATGAAAAAGTTTTTAGAAGCCATTGCTCTCATCTCCAGAGACCATGCCAGAACACCTATGCAATGGTCGTGCGAAGAGCCAAATGCTGGCTTCTCTGGGCCCACCGGCAAGCCATGGTTCTACTTGAACGAATCTTTCAGAGAGGGAATCAATGTCGAAGACGAGCAAAAGGATCCAAACTCTGTGCTGGCTTTCTGGAAGGAAGCTTTGAGATTTAGAAAGGCACACAAGGATATTACTGTTTATGGCTATGATTTTGAGTTCATCGACCTGGACAATAAAAAGCTGTTTAGCTTTACCAAGAAGTACGAGAACAAGACTTTGTTTGCAGCCTTGAACTTTAGCTCTGATAACGTGGACTTTACGATTCCAGATAACAGTACCTCGTTCAAGTTGGAGTTTGGAAACTTCCCAAAGAAGGAGGTAGATGCCTCTTCCAGAACTTTGAAGCCATGGGAGGGTAGAATTTACGTATCTGAATGATTGACGATTGTGACAGATCGGAAGAGCAC

And the annotations indicate that the longest ORF is a determinate gene but in the same frame there is an ORF that has a stop codon and thus probably a truncated protein. Indeed the truncated protein has a lot of interest for me. 

The question is: How can I interpret these results, Does the second truncate ORF has significance or is it only an artifact?

[Brian Haas]

There could be a frameshift in there. You might load up the sequence into IGV, align the reads to it, and examine the read support along the position of the potential frameshift.  If it was an assembly error, the read alignments should indicate this.

Attached is a screenshot of the ncbi longorfs output, just for reference.  (from https://www.ncbi.nlm.nih.gov/orffinder/)

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Brian J. Haas
The Broad Institute
http://broadinstitute.org/~bhaas