Purpose:

In this lab we will begin auto-annotating the Link.fasta file using PECAAN. Using GeneMark.hmm prokaryotic, we will be able to predict the coding potential for genes. We will also be comparing new sequences with sequences of  the known or assumed using BLAST.

Procedure:

DNAMaster Stetup

• Open link in FASTA file
• Go to https://discover.kbrinsgd.org/autoannotate/ and upload Linkfasta A file and enter phage name
• Click “Process”
•  Select and Copy the entire contents of the text box
• click “documentation” on DNA Master and paste information
• Parse information with default settings
• Save genome to downloads

GeneMark

• Go to http://exon.gatech.edu/GeneMark/gmhmmp.cgi (Links to an external site.)Links to an external site.
• Upload Link.fasta file
• Select the species Anthrobacter_aurescnens_TC1
• Under “Options” check the boxes labeled “Protein sequence”,”Gene nucleotide sequence”, and “PDF”
• Open the PDF and observe the coding potential for each gene

BLAST

• Go to https://blast.ncbi.nlm.nih.gov/Blast.cgi
• select gene #2 from DNAMaster and copy and paste the information from the “product” tab into the space provided
•  click BLAST and select the closest phage match to our gene
• copy the name of the phage and put this in the “notes” of our DNAMaster

Auto-Annotation

• Copy the template saved on DNAMaster from the previous lab into the notes section
• Fill in template based on  information given on ORFS and RBS of gene #2

Results

Raw SD score: -2.839

Generic Z Value:- 2.641

Final Score:-4.441

Start Codon:345

Stop Codon:560

There was a 4bp overlap

Conclusions

We found  a solid match to our gene through the NCBI protien BLAST database named Anthrobacter_aurescnens_TC1.  The final score on the Annotation was -4.441, which is good because it is the closest to zero.