Rationale: 

The purpose of today’s lab was to finish annotating the final 2 genes assigned in NapoleonB

Tools:

• DNA Master
• NCBI BLASTp
• HHPred
• Phamerator
• Starterator
• PhagesDB BLASTp
• Genemark (pdf)

Procedure:

• Lab began immediately with finishing up final annotations of assigned genes from the previous lab.
• Ran genes 63 and 64 through a series of BLASTp searches in both NCBI and PhagesDB, searched for protein folding structure in HHpred, analyzed the starterator analysis through PhagesDB, and checked for any supporting information of synteny by examining similar phages in the AM cluster with NapoleonB.

Results:

• Final Annotation for Gene 63:
  • SSC:41221 – 41427, CP:Yes, SCS:BothGM, ST:SS, BLAST-Start:Aligns with Arthrobacter Phage Arcadia gp62 NCBI BLAST q1:s1 0.95 2E-32, Aligns with Nason gp62 PhagesDB BLAST q1:s1 0.96 1E-28, Gap:4bp overlap, LO:NA, RBS:Kibbler7 and Karlin Medium 1.104 -6.698 Yes, F:NKF, SIF-BLAST:NKF, SIF-HHPred:NKF supported by gp , SIF-Syn:NKF
• NCBI BLASTp Results:
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• PhagesDB BLASTp Results:
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• HHPred Results:
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• Phamerator Results:
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• Final Annotation for Gene 64:
  • SSC:41417 – 41782, CP:Yes, SCS:Both, ST:SS, BLAST-Start:Aligns with Arhtrobacter Phage Correa gp60 NCBI BLAST q1:s1 0.64 1E-46, Aligns with Arthrobacter Phage Tribby gp65 PhagesDB BLAST q1:s1 0.76 1E-38, Gap:9bp overlap, LO:NA, RBS:Kibbler7 and Karlin Medium 3.16 -2.354 No, F:NKF, SIF-BLAST:NKF, SIF-HHPred:NKF supported by gp , SIF-Syn:NKF
• NCBI BLASTp Results:
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• PhagesDB Results:
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• HHPred Results:
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• Phamerator Results:
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Conclusions:

For both genes, the protein call was decided to be NKF as there was not enough supporting information to call a protein function with confidence. Despite there being a potential hit on HHPred, the probability was not enough to be sufficient for a call. Also the RBS for gene 63 despite having a low z-score was chosen as other suggested RBS shortened too much of the gene.

Next Steps:

The next steps for this lab are to edit and finalize any errors in the gene annotations made.