April 12

Progress Report and Continued Research 4/10/19

Progress Report and Continued Research 4/10/19

Rationale

The rationale behind today in lab was to search the literature to have us submit a progress report so that we and our coaches could see where we are at and what still needs to be done. In addition, when we were done with our progress report, we continued working on sequencing and protein folding in order to collect the necessary data.

Tools/Procedure

  1. Progress report was submitted
  2. Clustal Omega was used to compare TMPs between individual Arthrobacter clusters and between different tail morphologies
    1. Sipho and myo family groups were compared
  3. MEME Motif was used to find repeating motifs in the TMPs of sipho and myo phage

Results

The results above were generated using the MEME software to find motifs, areas of similar sequences across several different sequences belonging to both sipho and myo phage. As can be seen in the motif location image, there are two motifs (shown in blue and in green) that are present in every TMP tested, creating intriguing information to look into.

Conclusion

While there is not enough data to make extensive conclusions yet, our data so far suggests that TMPs are highly conserved (based on amino acid sequences) in each cluster, and less conserved in the tail families. The amino acid sequences do however suggest certain structural elements being conserved among tail families even if the amino acid sequence is not strictly conserved. Also, protein folding between myo and sipho phage showed significant differences in the protein structure, which we will need to examine further. Finally the results that show there are certain motifs found in every TMP seem to suggest areas of conservation.

Future Plans

In the future, we will continue to look for similarities and difference in the amino acid while continuing our folding of proteins to try to figure out if there are key conserved regions in TMPs.

Category: Lucy FIsher, Uncategorized | LEAVE A COMMENT
April 12

Literature Search Report 4/8/19

Literature Search Report 4/8/19

Rationale

The rationale behind today in lab was to search the literature to help us better understand what we are researching and to see if we can find valuable insights. In addition, when we were done with our literature search, we continued working on sequencing and protein folding in order to collect the necessary data.

Tools/Procedure

  1. Five sources of primary literature were cited and submitted
  2. Clustal Omega was used to compare TMPs between individual Arthrobacter clusters and between different tail morphologies
    1. AK, AL, AM, and all F clusters were compared

Results

The clustal omega results shown above help illustrate the results that were found during this lab period. While there is an incredible similarity between TMPs of the same cluster, even TMPs from the same tail morphology, but different clusters show fairly significant differences in the amino acid sequences. However, there does seem to be more similarity between the types of amino acids (as seen by the color coding) suggesting that even if DNA or amino acid sequences are not highly conserved, the structure is more likely to be conserved.

Conclusion

While there is not enough data to make extensive conclusions yet, our data so far suggests that TMPs are highly conserved (based on amino acid sequences) in each cluster, and less conserved in the tail families. The amino acid sequences do however suggest certain structural elements being conserved even if the amino acid sequence is not strictly conserved.

Future Plans

In the future, we will continue to look for similarities and difference in the amino acid while continuing our folding of proteins to try to figure out if there are key conserved regions in TMPs.

Category: Lucy FIsher, Uncategorized | LEAVE A COMMENT
April 12

April 8th + 10th- labs

  • APRIL 8TH 2019
  • OBJECTIVE: 
    • Find more primary literature to read in order to deepen our understanding of the project being worked on 
  • PROCEDURE: 
    • NCBI was used to search for sources 
    • Annotations were then created for those sources 
  • RESULTS: 
    • Sources listed below:
    • https://academic.oup.com/femsre/article/30/3/321/546048
    • Weigel, Christoph, and Harald Seitz. “Bacteriophage Replication Modules.” FEMS Microbiology Reviews 30, no. 3 (May 1, 2006): 321–81. https://doi.org/10.1111/j.1574-6976.2006.00015.x.
    • https://www.sciencedirect.com/science/article/pii/S1084952117306055
    • Hernandez, Alfredo J., and Charles C. Richardson. “Gp2.5, the Multifunctional Bacteriophage T7 Single-Stranded DNA Binding Protein.” Seminars in Cell & Developmental Biology, SI: Human dendritic cells, 86 (February 1, 2019): 92–101. https://doi.org/10.1016/j.semcdb.2018.03.018.
    • https://doi.org/10.1016/S0378-1097(99)00068-3
    • Hamann, Christian, Jörg Hegemann, and Armin Hildebrandt. “Detection of Polycyclic Aromatic Hydrocarbon Degradation Genes in Different Soil Bacteria by Polymerase Chain Reaction and DNA Hybridization.” FEMS Microbiology Letters 173, no. 1 (April 1, 1999): 255–63. https://doi.org/10.1111/j.1574-6968.1999.tb13510.x.
    • https://academic.oup.com/mbe/article/23/9/1688/1014265
    • Filée, Jonathan, Eric Bapteste, Edward Susko, and H. M. Krisch. “A Selective Barrier to Horizontal Gene Transfer in the T4-Type Bacteriophages That Has Preserved a Core Genome with the Viral Replication and Structural Genes.” Molecular Biology and Evolution 23, no. 9 (September 1, 2006): 1688–96. https://doi.org/10.1093/molbev/msl036.
    • https://www.sciencedirect.com/science/article/pii/S0042682218300588?via%3Dihub
    • Jalasvuori, Matti, and Katariina Koskinen. “Extending the Hosts of Tectiviridae into Four Additional Genera of Gram-Positive Bacteria and More Diverse Bacillus Species.” Virology 518 (May 1, 2018): 136–42. https://doi.org/10.1016/j.virol.2018.02.014.
  • CONCLSUION: 
    • Articles were read, and annotations were made 
  • FUTURE STEPS: 
    • Continue researching gene functions
  • –APRIL 10TH 2019
  • OBJECTIVE: 
    • Create a document with all the sequences that need to be run through data bases 
  • PROCEDURE:
    • A document was created and the gene for the terminase of a phage belonging to each of the following clusters was created:
    • AU1 AU2 AU3 AW BI1(bing) BI2 BI3 BI4 CC DJ EL  
  • RESULT:
    • N/A
  • CONCLUSION: 
    • Document was created 
  • FUTURE STEPS: 
    • Finish adding codes for each terminase protein for a phage of each cluster 
Category: Lauren Foley | LEAVE A COMMENT
April 12

4/10 Lab Journal

Purpose: Continue Individual Research

Procedure:

  • Searched for Phages containing an NMT in their genome to compare to the NMT found in NapoleonB and other AM cluster phages
  • Found three AU cluster phages for future research
  • Found three EG cluster phages with NMT genes
  • Added all the fasta files for the NMT genes from the new phages, AM cluster, and bacteria to a splitstree diagram
  • Fought splitstree for almost a half hour to get it to work
  • Eventually manipulated the program to do its job
  • Found that with the present data we couldn’t put together a map to truly represent the genealogy of the gene

Results: Found that the splitstree graph will be harder to create than previously imagined. Will need more samples.

Future Plans: Continue finding more NMT fasta files to make into a coherent phylogenetic tree

Category: Uncategorized | LEAVE A COMMENT
April 12

4-10-19 — Data Gathering – AQ and AR Clusters

Data Gathering – AQ and AR Clusters

Date: 4-10-19

  • Rationale
    • The rational for this lab is to continue gathering data to use for our independent research project.
  • Procedure
    1. PhagesDB was used to find the entire nucleotide sequence for phages in the AQ and AR clusters.
    2. SplitsTree was used to produce phylogenetic trees for the AQ and AR cluster phages.
    3. Start codon preferences for each phage were calculated and recorded on the google sheets page used in previous labs.
  • Results
    • Above is a screenshot of a phylogenetic tree including every AR cluster phage. This was made using whole genome alignment between the phages.
    • Above is a screenshot of a phylogenetic tree including every AQ cluster phage. This was made using whole genome alignment between the phages.

  • The preferred start codons of phage KBurrousTX

    The preferred start codons of phage Linus

  • Future Plans
    • The next step is to continue gathering this data before attempting to further subcluster phages based on start codon preference.
Category: Brandon Reider | LEAVE A COMMENT
April 12

4-8-19 — Data Gathering

Data Gathering

Date: 4-8-19

  • Rationale
    • The rational for this lab is to continue gathering data to use for our independent research project.
  • Procedure
    1. PhagesDB was used to find the entire nucleotide sequence for phages in the AM cluster.
    2. SplitsTree was used to produce a phylogenetic tree for the AM cluster phages.
    3. Start codon preferences for each phage were calculated and recorded on the google sheets page used in previous labs.
  • Results
    • Above is a screenshot of a phylogenetic tree including every AM cluster phage. This was made using whole genome alignment between the phages.

  • NapoleonB’s preferred percentages of each start codon

    Mudcat’s preferred start codon of each gene

  • Future Plans
    • The next step is to continue gathering this data before attempting to further subcluster phages based on start codon preference.
Category: Brandon Reider | LEAVE A COMMENT
April 12

Independent Research Project 4/10/19

Rationale

Today we will continue working on our independent research projects and analyze the data that we have collected.

Procedure

  • Two-sample t-tests were performed on the data previously collected.
  • Pham numbers were changed since data on PhagesDB was updated. We spent the remainder of lab time double-checking to ensure that the same genes there were declared as AM specific before remained AM specific.
  • Our groups were altered and our results were changed.

Results

The computed averages are shown below.

Conclusions/Next Steps

Next, we aim to collect more data on AM phages and non-AM cluster phages.

Category: Emily Gaw | LEAVE A COMMENT
April 12

Protein Prediciton 4/10/2019

Title: Protein Prediction

Date: 10 April 2019

Rationale: With the discovery of the pham abnormality, further inspection will be done on the proteins to see if any more information can be found on the proteins and their relation to the sequence.

Procedure: The proteins were BLASTed and loaded into HHpred in order to inspect the possible matches or structure of the proteins. Further inspection was done on surrounding sequences and proteins in order to possibly discover more about the sequence of interest.

Results/Observations: There were no significant BLAST hits for the proteins, but more research will be done to discover the nature of the proteins.

Conclusions/Next Steps: More tools like RaptorX or other protein structuring tools will be utilized to possibly draw comparisons between proteins that could show a reasoning for the sequence of interest.

Category: Cooper Johnson | LEAVE A COMMENT
April 12

Independent Research Project 4/8/19

Rationale

Today we will continue working on our independent research projects. We aim to gather more data on NapoleonB and other AM cluster phages.

Procedure

  • The annotations created by the class were uploaded onto DNA Master.
  • The %GC and %GC3 were calculated for each gene in Napoleon and the results were recorded into an excel sheet.
  • The genes and their %GC and %GC3 values were sorted into two categories of whether the gene was AM specific or non-AM specific.
  • The average %GC and %GC3 values were calculated for each group created.
  • The process was repeated for a non-AM cluster phage.
  • Average %GC was also collected for each specific type of Arthrobacter phage.

Results

Shown below are the recorded average %GC for each cluster of Arthrobacter phage.

Conclusion/Next Steps

Next, we will perform a t-test with our results and move onto data collection for other Arthrobacter phages.

Category: Emily Gaw | LEAVE A COMMENT
April 12

Protein Structures of the AM Cluster (4/10/19)

Rationale: Although some of the data points have not been entered for the tape measure protein, due to the time frame, protein structures for the major tail protein in the AM cluster were looked at.

 

Tools:

  • Personal Computational Device
  • Google Excel
  • PhagesDB
  • Swiss-Model

 

Results:

  • With Google Excel, the major tape protein amino acid sequences in the AM cluster were each individually copied and pasted into the Swiss-Model program which then generated a model for the protein.
  • These models were screenshotted and saved in separate files to ensure that the data remains organized.

Heisenberger Major Tail 15 AUG

 

Heisenberger Major Tail 15 GUG

 

Heisenberger Major Tail 15 UUG

Conclusion

  • The protein structures from the AM cluster were looked at and organized to avoid confusion.

 

Future Plans:

  • Protein structures will be continued to be observed and organized.
Category: Sabin Patel | LEAVE A COMMENT