Rationale:

The purpose of today’s lab was to go over the use of Phamerater and tRNA searching to add on to the toolkit used for annotation. Also genes were assigned for us to check to further practice our annotation skills.

Tools:

• DNA Master
• Phamerator
• Aragorn
• PhageNotes

Procedure:

• Aragorn and tRNAscan-SE were introduced to practice searching for tRNA folding in phage annotation.
• Phamerator accounts were set up to compare annotated genomes of phages in the same cluster as phage Elesar.
• Genes 30 and 31 were revised by checking the Starterator and Synteny regions of annotation notes.
• Was assigned genes 4 and 5 to check over to see if annotations were correct, so genes were annotated fully to check for any errors.

Results:

• Final Annotation Notes for Gene 4:
  • SSC:1349, 1984, CP:Yes, SCS:Both, ST:SS, BLAST-Start:Aligns with Nandita gp5 NCBI BLAST q1:s1 0.95 3E-122, Aligns with Nandita gp5 PhagesDB BLAST q1:s1 0.95 3E-122, Gap:overlap 4, LO:, RBS:Kibbler7 and Karlin Medium 2.23, -4.085 ,no, F:terminase, small subunit, SIF-BLAST:terminase small subunit Supported by NCBI BLAST Nandita gp5 AYN58627 0.95 3E-122, terminase small subunit Supported by PhagesDB BLAST Nandita gp5 0.95 3E-122, supported by a CD found from the databse gp , , SIF-HHPred:terminase small subunit, COG, prophage, gpNA, COG3747, 0.75, 99.89, SIF-Syn:
  • 
  • 
• Final Annotation Notes for Gene 5:
  • SSC:1971 ,3668, CP:Yes SCS:Both, ST:SS, BLAST-Start:Nandita, gp6, NCBI BLAST, q4:s5 0.99 0, Aligns with Nandita gp6 PhagesDB BLAST q4:s5 0.99 0, Gap:13bp overlap, LO:Yes, RBS:Kibbler7 and Karlin Medium 1.724 -5.684 No, F:terminase, large subunit, SIF-BLAST:terminase large subunit Supported by NCBI BLAST Nandita gp6 AYN58628 0.99 0, terminase large subunit Supported by PhagesDB BLAST Nandita gp6 0.99 0, supported by a CD found from the databse gp , , SIF-HHPred:terminase large subunit supported by Terminase_1 gpNA PF03354.15 0.87 99.8, SIF-Syn: 
  • 

Conclusions:

The main proteins called by both NCBI BLASTp and HHpred seemed to call both small and large subunit phage terminases in genes 4 and 5 respectively. It can be assumed based off the supporting data that these are the proteins coded by genes 4 and 5 as both probabilities were above 90 percent in both cases.

Next Steps:

The next steps for this experiment are to practice searching for synteny within the genome and also practice further with annotating genes. Now that phamerator have been introduced, gene comparison between phage Elesar and similar phages in the cluster.