Tutorial

Tutorial for Prophage Hunter

1. To launch Prophage Hunter, click the “Start Hunting” button at the navigation bar of Prophage Hunter web page (Fig. 1).

Click the button in the red box to launch HAPp

Fig. 1. Click the button in the red box to launch Prophage Hunter.

2. On the main page, click “Browse” to upload one or more nucleotide sequence(s) in a FASTA file (Fig. 2a). By default, Prophage Hunter uses a similarity search strategy to identify the initial prophage region. Alternatively, by ticking the “Skip similarity matching” box (Fig. 2b), users can omit this process in order to identify novel phages.

Fig. 2. Click “Browse” to upload FASTA file.

3. Enter your email address to receive the hyperlink directed to the analysis report (Fig. 3). Please note that this hyperlink will be expired ONE WEEK later.

Fig. 3. Enter email address to receive the analysis report.

4. Tick the box to join “User Experience Improvement Program” (Fig. 4a). For more details click “User Experience Improvement Program” (Fig. 4b).

Fig. 4a. Enter email address to receive the analysis report.

Fig. 4b. User Experience Improvement Program.

5. Click “START HUNTING” button to start your analysis.

Click “START HUNTING” to search active prophages

Fig. 5. Click “START HUNTING” to search for active prophages.

6. Wait until the analysis is completed (Fig. 6).

Fig. 6. Wait until your analysis is completed.

7. On top of the main output is a genome browser showing each predicted prophage region (Fig. 7). Active and ambiguous regions are colored in azure and gray, respectively (Fig. 7a). Users can switch to other genomic DNA sequences by clicking the dropdown menu on the top left (Fig. 7b).

Fig. 7. A genome browser showing locus and category of each predicted prophage region.

8. On the bottom is a table showing details of each predicted prophage region (Fig. 8a):

- Candidate ID: ID of the predicted prophage region;

- Sequence ID: ID of the genomic sequence shown in the input file;

- Start: start position of the predicted prophage region;

- End: end position of the predicted prophage region;

- Length: length of the predicted prophage region;

- Category: the category of the predicted prophage region;

- Score: score of the predicted prophage region, the higher the score is, the stronger the evidence that supports an active prophage region;

- Closest phage: the closest phage of the predicted prophage region compared to known phages, “N/A” represents not homologous to known phages;

- Gene number: number of predicted prophage genes within the predicted prophage region.

Fig. 8. Details of each predicted prophage region.

Users can click the corresponding buttons to download genomic DNA sequences of each region, or CDS of predicted prophage genes, or amino acids sequences of predicted prophage genes (Fig. 8b), or all analysis results (Fig. 8c). Users can also click the taxonomy name of the closest phage to see the details (Fig. 8d).

9. Click the predicted prophage region in genome browser or table to see annotation of each gene (top, Fig. 9a) and top 5 closest phages that homologous to this region (bottom, Fig. 9b).

Fig. 9a. NCBI_nr, Pfam and InterPro annotation of each predicted prophage gene within this region.

Fig. 9b. Top 5 closest phages that homologous to this region.