Ashton Peckinpah

Tuesday, February 27, 2017

Objective: To annotate the entire Timinator genome by having each student annotate five genes. This allowed for each student to feel more comfortable annotating genes as we approach the beginning of our research projects. Today, I was given the challenge to annotate assigned genes for Timinator. I was assigned:13, 35, 57, 80, and 68.

(Tools Used): DNA Mastering, GeneMark, Phagesdb, Starterator, Phamerator, Google Docs, and HHPred

Methods: First, I had to run an auto-annotation of the Timinator draft in DNA master. For each gene assigned, I had to complete each of the following portions of the annotation:

• SSC:/ Gap or overlap: start/ stop and gap/ overlap distance.
• CP: covered by all.
• SCS: Does it agree with Glimmer/ GeneMark.
• NCBI Blast: Best blast hit, E-Value, Conserved Domain hit, and CDD E-value.
• SD: SD score, ranking, and Z-value.
• PhagesDB Blast: Best blast hit, E-Value.
• HHPred: Best blast hit, E-value.
• LO: longest open reading frame.
• ST: does it agree with Starterator.
• F:/ FS: function and support for this function.

After completing the above portions of the annotation I  inserted the annotation components into the Timinator Google document. Every time I had to update the status on the home page and confirm that the gene annotation was/is complete.

Results: 

Gene 13-

Start: 9980bp Stop: 10591bp FWD GAP: 0bp Gap SD Final Value: SD Score: -3.467 (Best score) Z-Value: 2.701 CP: The gene is covered SCS: Agrees with Glimmer, Agrees with GeneMark NCBI BLAST: hypothetical protein Arthrobacter phage BarretLemon_13 E-Value: 0.0 CDD: No good hit PhagesDB BLAST: DarthPhader_8, function unknown, 307 E-Value: 2e-10 HHPred: No good hit LO: Yes ST: Agrees with Starterator F: NKF FS: NCBI, PhagesDB Notes:

(Phagesdb)

(RBS table results)

Gene 35-

Start: 29826bp Stop: 30671bp FWD GAP: 2bp Gap SD Final Value: SD Score: -2.275 (Best score) Z-Value: 3.294 CP: The gene is covered SCS: Agrees with Glimmer, Agrees with GeneMark NCBI BLAST: RecT-like recombinase [Arthrobacter phage BarretLemon] E-Value: 0.0 CDD: RecT Superfamily E-Value: 9.89e-98 PhagesDB BLAST: StevieBAY_Draft_35, function unknown, 281 E-Value: e-161 HHPred: No good hit LO: Yes ST: Agrees with Starterator F: NKF FS: Phages DB Notes: NCBI said the function of this gene was a RecT DNA recombinase, but this function option was an option in PhageNotes.

(NCBI results) 

(C0nserved Domain)

(Phages db)

Gene 57-

Start: 40263bp Stop: 40526bp FWD GAP: 4bp Overlap SD Final Value: SD Score: -6.513 (Best score) Z-Value: 1.478 CP: The gene is covered SCS: Agrees with Glimmer, Agrees with GeneMark NCBI BLAST: HTH DNA binding protein [Arthrobacter phage BarretLemon] E-Value: 8e-42 CDD: No good hit PhagesDB BLAST: StevieBAY_Draft_57, function unknown, 87 E-Value: 3e-42 HHPred: No good hit LO: Yes ST: Agrees with Starterator F: HTH DNA binding protein, MerR-like FS: NCBI Notes:

(Phagesdb)

(RBS results)

Gene 68-

Start: 43977bp Stop: 44168bp FWD GAP: 4bp Overlap SD Final Value: SD Score: -3.697 (Best score) Z-Value: 2.625 CP: The gene is covered SCS: Agrees with Glimmer, Agrees with GeneMark NCBI BLAST: hypothetical protein BARRETLEMON_67 [Arthrobacter phage BarretLemon] E-Value: 8e-38 CDD: No good hit PhagesDB BLAST: LeeroyJ_Draft_68, function unknown, 76 E-Value: 2e-36 HHPred: No good hit LO: No Better SD score ST: F: NKF FS: NCBI, PhagesDB Notes:

(Phagesdb)

(RBS Result)

Gene 80-

Start: 50855bp Stop: 51088bp BKWD GAP: bp Gap SD Final Value: SD Score: -6.658 (Best score) Z-Value: CP: The gene is covered SCS: Disagrees with Glimmer, Agrees with GeneMark Genemark gave me a better SD score NCBI BLAST: hypothetical protein BARRETLEMON_79 [Arthrobacter phage BarretLemon] E-Value: 1e-47 CDD: No good hit PhagesDB BLAST: StevieBAY_Draft_79, function unknown, 77 E-Value: 1e-39 HHPred: No good hit LO: Yes ST: F: NKF FS: NCBI Blast, PhagesDB, HHpred Notes:

This gene was deleted (per professor) since it had a 100% overlap with gene 79.

Additionally, the gene had no known function.

In conclusion: We were able to put all of our semester work to the test. As a class, we successfully annotated the entire genome of Timinator. Now, it can be sent off to Pittsburg to be approved.  This annotation will be useful in the future when researching gene functions, promoters, and other parts of the genome.