Database practice

Explore a gene

SopD is an effector protein of Salmonella Typhimurium functions to alter host cell physiology and promote bacterial survival in host tissues. It contributes to the replication of bacterial in macrophages. We will explore the detail of sopD gene in multiple databases in order to understand what do we expect from these databases in Bioinformatic study.

NCBI GenBank

  1. Go to http://www.ncbi.nlm.nih.gov/ , enter salmonella typhimurium[orgn] AND sopd and click Search. By default you search ALL the databases and the results also include many that refer to this topic but are not necessarily specifically about sopD. How many nucleotide sequences are returned? ___

    Click at ‘Nucleotide’ to see the Result Page. Examine the accession number for each entry in the new page.

  2. Click on the sequence “Acession: AF234265.1”.

    1. Read the GenBank file. When did the sequence uploaded? __ How many publications are there about this sequence? __
    2. Click at Graphics in the top left. Explore the graph, Try zoom in, zoom out, and move left or right. What’s the start and end position of the gene on the Salmonella Typhimutium genome? __ to __
    3. Click at Fasta in the top left, and then click at send to in the top right, choose file, save the file to a local folder as sopD_gene.fasta

UniProt

  1. Go to https://www.uniprot.org/, enter salmonella typhimurium sopd and click Search. How many manually reviewed results, and how many unreviewed results are returned? ___
  2. Click on the Entry P40722. Read the results. Try find the following information:
    1. What’s the subcellular location of the protein?
    2. What’s the resolution of the protein structure?
    3. Download the fasta file and save as sopD_protein.fasta.
    4. Find the KEGG link from the Genome annotation databases. Go to KEGG

KEGG

  1. Read the information on the KEGG page.
  2. Click the link after Pathway. What’s the downstream protein of sopD in the “Bacterial invasion of epithelial cells” pathway?

Download a reference genome

  1. Go to http://www.ncbi.nlm.nih.gov/ and enter salmonella typhimurium and click Search. How many results are returned from assembly database?
  2. Click on assembly database, choose the entry ASM694v2.
    1. Read the page, click the link after Organism name. Which journal and when did the sequence first published?
    2. Back to the ASM694v2 page, click the Download Assembly button at the top right. Download the genomic fasta, genomic GFF and genomic genbank file from refseq database. Save as salmonella_typhimurium_lt2.fasta, salmonella_typhimurium_lt2.gff and salmonella_typhimurium_lt2.gb
  3. Back to the NCBI search result page, this time go to SRA database.
    1. Choose the first entry. Now you see the record of a sequencing data of a Salmonella Typhimurium strain. What’s the sequencing machine used? What’s the size of the raw data?
    2. Click the link after Sample. This is the metadata of the Salmonella Typhimurium strain bacterial. Where, when and from which source did this bacterial sampled?
    3. Back to the SRA page. Click the link under run, now you will see the metadata of the sequence.
    4. Open terminal, login to the server, download the sequence use sra-tools with the SRR run id:
     source activate bioinfo
 fastq-dump --split-3 --gzip SRR10561173

    It may take a long time. Be patient, we don’t need this file today.

    More about sra-tools: sra-tools documentation

Visualise the reference genome

  1. Download the Artemis installation package and install.
  2. Start Artemis, click File - Open, open salmonella_typhimurium_lt2.fasta, load the genome sequence
  3. Click File - Read an entry, open Salmonella_typhimurium_lt2.gff, load the annotation file.
  4. Getting around in Artemis
  5. Try the Goto menu. Click Goto - Navigator
    1. Fill a number in goto base form. Then click Goto
    2. Fill sopD in Goto Feature with gene name, then click Goto
  6. Try the Select menu. Click Select - Feature selector, Select and view all tRNAs
  7. Take your time play with Artemis!