4/19 Emily Johnson

Purpose: The goal for today was to use bioinformatics tools to compare our HNH Endonuclease sequences

Materials:

TM-Align

SWISS-MODEL

Methods:

We divided up the work today: Navya was in charge of using Phamerator to gather data about our gene, such as GC content, pham numbers, etc. Alex looked up info about HNH Endonucleases, and I used TM-Align and SWISS-MODEL to compare two protein structures at once.

Results:

Purpose: The purpose of lab today is to work on our poster for the URSA Scholar’s Week

Methods: Today we divided up the tasks we need to do for the poster. I am in charge of doing the introduction and materials. Navya is in charge of the wet lab and in silico methods. Josh will be explaining the results and conclusion and Thomas will be responsible for taking all of our materials and designing the poster for our finished product.

I was able to make a list of all the materials we needed today, and work on part of the intro. I used an article or two to describe the importance of arthrobacter phages and had to ask around some other groups to help me remember exactly what we did in the first semester. Also, I had to do some research for what the uses of arthrobacter are.

Next lab, we will be presenting the poster, so I will finish my introduction tonight and send it Thomas so he can put it together.

Data:

GeneMark Coding potential, Gene 23 is the second dark line on the third reading frame, Gene 24 is the dark line right after the small white gap following Gene 23, also on the third reading frame. Gene 25 begins on the dark line on the right side of the first reading frame.

Gene 23:

NCBI Hit Gene 23

Phages DB Hit Gene 23

Gene 24:

NCBI Hit Gene 24

Phages DB Hit Gene 24

Gene 25:

HHPred Hit Gene 25

CDD Hit Gene 25

NCBI Hit Gene 25

Phages DB Hit Gene 25

Results:

Gene 23:

Start: 17661bp Stop: 18473bp FWD GAP: 171bp Gap SD Final Value: SD Score: -2.589 (2nd best score) The first best score causes a big gap that is uncharacteristic of bacteriophage genomes and there is no potential for a gene between the two. Z-Value: 2.923 CP: The gene is covered SCS: Agrees with Glimmer, Agrees with GeneMark NCBI BLAST: hypothetical protein LAROYE_24 [Arthrobacter phage Laroye] Q1 S1 E-Value: 1e-112 CDD: No good hit PhagesDB BLAST: Laroye_24, function unknown, 280 Q1 S1 E-Value: 1e-112 HHPred: No good hit LO: Yes ST: F: FS: Notes:

Gene 24:

Start: 18477bp Stop: 18854bp FWD GAP: 3bp Gap SD Final Value: SD Score: -3.779 (Best score) Z-Value: 2.852 CP: The gene is covered SCS: Agrees with Glimmer, Agrees with GeneMark NCBI BLAST: hypothetical protein LAROYE_25 [Arthrobacter phage Laroye] E-Value: 8e-62 CDD: No good hit PhagesDB BLAST: Wheelbite_Draft_23, function unknown, 127 Q1 S1 E-Value: 9e-51 HHPred: No good hit LO: Yes ST: Agrees with Starterator F: NKF FS: Notes: This gene surprised me a bit because it starts 3 base pairs after the prior gene, which would not be weird except that they are on the saem reading frame so it almost looks like it could be one long gene save for the stop codon between them.

Gene 25:

Start: 18889bp Stop: 19848bp FWD GAP: 34bp Gap SD Final Value: SD Score: -2.293 (Best score) Z-Value: 3.068 CP: The gene is covered SCS: Agrees with Glimmer, Agrees with GeneMark NCBI BLAST: capsid protein [Arthrobacter phage Laroye] Q1 S1 E-Value: 0.0 CDD: Phage_cap_E pfam03864 E-Value: 1.29e-3 PhagesDB BLAST: Salgado_26, capsid, 319 Q1 S1 E-Value: 1e-174 HHPred: Phage-related protein; structural genomics, joint center for structural genomics, JCSG, protein structure E-Value: 4e-40 LO: Yes ST: Agrees with Starterator F: major capsid protein FS: NCBI, PhagesDB, and HHPred all call it a major capsid protein Notes:

Data:

Gene 20:

NCBI Blast results for gene 20

PhagesDB Blast results gene 20

Gene 21:

NCBI Blast result gene 21

PhagesDB Blast result gene 21

Gene 22:

NCBI Blast result gene 22

CDD Hit Gene 22

PhagesDB Hit Gene 22

HHPred Hit Gene 22

Results:

Gene 20:

Start: 16017bp Stop: 16466bp FWD GAP: 79bp Gap SD Final Value: SD Score: -3.108 (Best score) Z-Value: 3.074 CP: The gene is covered SCS: Agrees with Glimmer, Agrees with GeneMark NCBI BLAST: hypothetical protein LAROYE_20 [Arthrobacter phage Laroye] Q1 S1 E-Value: 9e-28 CDD: No good hit PhagesDB BLAST: Edmundo_Draft_19, function unknown, 149 Q1 S1 E-Value: 2e-71 HHPred: No good hit LO: Yes ST: Agrees with Starterator F: FS: Notes:

Gene 21:

Start: 16456bp Stop: 16863bp FWD GAP: 11bp Overlap SD Final Value: SD Score: -4.651 (2nd best score) The best score has a gap of 356bp and the gene would only be 42bp long Z-Value: 1.936 CP: The gene is covered SCS: with Glimmer, Agrees with GeneMark NCBI BLAST: hypothetical protein LAROYE_22 [Arthrobacter phage Laroye] Q11 S5 E-Value: 8e-49 CDD: No good hit PhagesDB BLAST: Salgado_22, function unknown, 122 Q11 S5 E-Value: 2e-46 HHPred: No good hit LO: No The longest reading frame caused a big overlap, bad Z and final scores and bad e values on hits from NCBI and PhagesDB ST: Agrees with Starterator F: NKF FS: Notes: In both NCBI and PhagesDB Blasts, the Query was 11 and the Subject was 5. I tried blasting this gene using the start codon before nad the start codon after, but this just created larger discrepancies between the Query and Subject matches, as well as unlikely overlaps, gaps, and Final/Z scores.

Gene 22:

Start: 16860bp Stop: 17489bp FWD GAP: 4bp Overlap SD Final Value: SD Score: -5.23 (3rd best score) The first two best scores cause a gap of at least 800 bps Z-Value: 1.695 CP: The gene is covered SCS: with Glimmer, Agrees with GeneMark NCBI BLAST: No good hit CDD: Rnase_HI_RT_non_LTR cd09276 E-Value: 2.43e-09 PhagesDB BLAST: Salgado_23, RNAse, 209 Q1 S1 E-Value: 1e-88 HHPred: Ribonuclease H1 E-Value: 2.4e-18 LO: Yes ST: Agrees with Starterator F: NKF FS: NBCI. PhagesDB and HHPred call it an RNAse (ribonuclease) Notes: The function is actually predicted to be a RNAse but that is not an option on the function list