Theresa WangChin Wong

Date of Lab: April 18

Tools used: SwissModel, PhagesDB, DNAmaster, RaptorX

Procedures:

• We got all our results back from RaptorX.
• We decided we will only use the results from RaptorX instead of SwissModel due to limited space on the poster, but there was not much difference between raptorX and SwissModel.
• We found the percentage of similarity between Timinator and each of the seven phages
• To help explain the function of Holin, we reasearched a figure with details of how the Holin protein work on destroying the membrane

Conclusion:

In the future we hope to find out how is Holin protein related to some diseases and how will understanding the function of protein help with cueing the diseases.

Theresa WangChin Wong

Date of Lab: April 11

Tools used: SwissModel, PhagesDB,DNAmaster, RaptorX

Procedures:

1. Download the finished DNAmaster file
2. Based on the gene number of the phages we recorded from last time, locate the Holin protein from each phages.
3. Record all the Product sequence of hollin genes of the phages we have chosen.
4. Copy and paste the protein sequences into SwissModel to generate a model of the structure of gene with Holin protein.
5. Sumbit job on RaptorX structure prediction server with the protein sequences for a more accurate results.
6. Start working on the abstract and outline of the poster.

Conclusion:

Considering that Swiss Model is just a prediction that match with known protein structures, we also submitted job on RaptorX, which can give us a more accurate model of the proteins. However, we will need to wait for RaptorX to send the result back to us, hopefully we will get all eight of the results back by next class.

Results that we got from SwissModel

Theresa WangChin Wong

Date of Lab: 03/28/17

Rationale: The purpose of the lab was to decide the main question and topic for the capstone project.

Tools used: phagesDB, asmblog , ncbi, SwissModel

Procedures:

We looked into the topic that we are interested in from last week, and we decided we want to pick the protein holin as our topic. We found out that holin protein exist in Timinator and Lore. We recorded the gene numbers of the holin protein and we found that there are proteins that are responsible for the lysis function of the bacterial membrane located right before and after holin protein. So we did more research on it and found out that holin protein are related to signaling the lysis protein to destroy the membrane.

We assume that holin exist in other phages as well, so we used Phamerator to find out other phages that are in the same pham as Timinator that has holin and we picked seven of those that have been annotated. Then we decided we want to know more about holin through the figures of the genes. We used Swiss Model to predict the structure of the gene and we found out that they all have similar structure.

Conclusion: We had much progress in this lab and learned more interesting facts about holin protein.

(Structure of Holin protein in Timinator from SwissModel)

Lab 10- Mar 21 – Project Day1

Date of Lab: 03/21/2017

Rationale:

We got into a group of three and our goal today is to find out the topic we are interested in for our project.

Tools used: phagesdb

I got into a group with Kiersten and Kirsty. With the help of phagesdb, we found out some topics we are interested in. We came up with some questions that we could possibly explore in the project and the rough outline of the poster for the project.

Methods: We can use the tools that we are already familiar with, like DNAmaster, Phamerator, Starterator and some software that can graph out the gene and allow us to compare the genes.
Conclusion:

The outline that we decided the poster to have is blue in color, white background, font size of 30, vertical boxes.

Date of Lab: 03/14/17

Rationale: The purpose of the lab was to continue to annotate our assigned gene of Timinator, come up with genomic questions for our final poster projects.

Tools used: DNAMaster, Phamerator, Starterator, GeneMark, NCBI Blast, CDD, HHPred

Procedures: Continue working on the gene annotation of Timinator (on gene 73 and 12)

Conclusion:

gene-12:hhpred-no good hit

gene12:phagesdb

Gene73:phagesdb

gene73-hhpred

Date of Lab: 02/28/17

Rationale: The purpose of the lab was to annotate 5 genes from Phage Timinator.

Tools used: DNAMaster, Phamerator, Starterator, GeneMark, NCBI Blast, CDD, HHPred

Procedures: Annotate Gene 21,43,65,73(B),12(B) through the tools listed above and record the results on spreadsheet.

Conclusion:

I only worked on gene 21,43 and part of 65 in this lab. I did not make any changes on gene21,43 and 65, they were all auto-annotated.

Gene 21: According to NCBI blast and Phamerator, it is a tail protein [Arthrobacter phage BarretLemon], so it has a function of major tail protein.

Gene 43 has no known function and has Q1S1.

Gene 65 has no known function and has Q1S1.

If there is no other starting site close to the auto annotated starting site and the gene was not fully covered(on geneMark), just keep the starting site unchanged even though it was not fully covered. If not it will create a huge overlap with the gene in front.

gene43

gene21

gene65 NCBI blast

gene65 HHpred

Date of Lab: 01/31/17

Rationale: The purpose of the lab was to practice genome annotation and learn how to use Phamerator.

Tools used: DNA master, NCBI, Phamerator

Procedures:

1. Open the software Phamerator through virtual box and login with the password “phamerator”.
2. Under “Phages” tab> search the interested gene (It will allow you to look at multiply genes at the same time) > MAP
   1. the number in the colored box = gene number
   2. number above the box = pham number
   3. number in (#) = number of members in that pham
   4. lines on the graph: identifying sequence that are similar on other part/ duplication/different place/ phage
   5. upper gene= forward
   6. lower gene = reverse
3. If we pointed at the gene/the rectangle, it would show the function of the gene(only apply on some genes)
4. Annotate the assigned gene (gene 22)

Conclusion:

We got to learn to use a new tool to help us to annotate the genes with more details and have more practice on annotating the genes.

Notes:

• each ruler = genome
• rectangle = auto annotated gene
• high gene density
  • shouldn’t have too many gap/overlap–should be packed
• there will be gap for promoter between genes
• If it is a perfect match :
  • query coverage ~200%
  • E value ~ 0
• Long DNA = tap measure gene
• Short gene = regulator
• F = function
• Fs = function source: blast, pharmarater, HHpred, cdd(conserved domine), record best blast

Theresa WangChin Wong

Date of Lab: 02/21/17

Rationale: The purpose of this lab was to present the annotation of our assigned and to ensure the annotation was accurate.

Procedures:

1. Add the Q:S value to the google spreadsheet.
2. Help reannotate the genes of other groups if necessary.

Notes:

*only shorten a gene If there’s too much overlap
*Enter new start site into DNA Master: input the new start site > hit the calculator button > “Post”

*When we are deciding the start site, we prefer to choose the gene with a better SD score than with the longer ORF.

Conclusion:

I learned a lot from learning about how the other gene area annotated and why changes were made on annotating the genes base on different situation.

Theresa WangChin Wong

Date of Lab: 02/14/17

Rationale: The purpose of this lab was to annotate Lore for the assigned gene and put the results on the excel spreadsheet.

Tools used: DNA master, NCBI, Phamerator, Starterator, Glimmer, GeneMark

Procedures:

• We did not make any changes on gene 13.
• Start: 7279bp Stop: 9276bp FWD GAP: 110bp Gap SD Final Value: SD Score: -4.389 (2nd best score) Z-Value: 2.209 CP: The gene is covered SCS: Agrees with Glimmer, Agrees with GeneMark NCBI BLAST: Tapemeasure protein Arthrobacterphage Toulouse E-Value: 0.0 CDD: Phage-related protein [Mobilome: prophages, transposons] E-Value: 5.90e-11 PhagesDB BLAST: StewieGriff_Draft_13, function unknown, 665 Q:1 S:1 E-Value: 0 HHPred: No good hit LO: Yes ST: Agrees with Starterator F: tape measure protein FS: Phages DB, NCBI, HHPred, Phamerator

• We chose the #1 gene with the 2^nd best score and the longest ORF to be our starting site. Since it also agrees with Glimmer, GeneMark and Starterator, and it cover most of the gene. We did not choose the gene with the best score(gene9) because it would create a larger gap.

• Gene #1 has the most number of blue vertical bars on Starterator and that’s the gene Starterator suggested.

We got a 0.0 E value on NCBI BLAST, its function is tapemeasure protein

• We got Query1: Subject 1 on NCBI BLAST, that’s mean our query gene match with the subject(top hit) gene with the same length. So we can agree with the start codon.

• Phagesdb: StewieGriff_Draft_13, function unknown, 665 has an e value of 0.0.
• Conserved domains
• NO hit on HHPred

Conclusion:

With the help of the excel spreadsheet, we did the annotation step by step and we are able to compare our results with other genes at the same time.

Theresa WangChin Wong

Date of Lab: 02/07/17

Rationale: The purpose of the lab was to annotate our assigned genes (ranging from 8401-9800) and learn how to use Starterator.

Tools used: DNA master, NCBI, Phamerator, Starterator, Glimmer, GeneMark

Procedures:

1. Download the Fasta file for Lore on fastdb website and load it into DNA master.
2. To figure out which gene we should annotate, we compare our assigned range with the 5’End & 3’End ranges (on the Features tab). And we found out we it mostly cover gene 13 and overlap with gene 14.
3. Fill in the start coordinate, stop coordinate, FWD/BKWD, Gap/Overlap
4. Coding potential :if the gene (black horizontal bar) is longer than the line graph, it is covered
5. Shine Delgarno :SD score & Z-Value
6. start choice source: does it agree with Glimmer & GeneMark?)
7. LO: longest ORF value
8. NCBI: blast it on NCBI, record the name of the best hit and its E-Value
9. Conserved Domains: Click the button above the line graph on the NCBI website. Record the best hit & its E-Value.

• Blast on Phages DB & HHPred
• Starterator: Open Virtue Box>Starterator>

”Choose what you want to Starterate” :Pham

“Pham Number”: 6177 find it on the Phamerator (the number on top of our assigned gene)

>Link to Report > see whether it agree with Starterator

start choice: base on the gene with the most blue vertical bar (other vertical bar with color than blue = random)

• Function: find it through Phamerator
• Function source: Phages DB, NCBI, HHPred, Phamerator

Conclusion:

I became more familiar with the whole process of annotating the gene, with a organized google spreadsheet. And the picture below is the result of my  annotation.