Rationale:

To understand more about phage, the class was divided into several small groups to generate research questions that utilize bioinformatic tools to learn more about questions about NapoleonB and/or related phages.

Materials:

• Laptop
• Canvas Bio-Lab info page
• DNA Master
• NCBI & Phagesdb Database
• Phire

Procedure & Results:

The NapoleonB genome was analyzed on DNA Master for promoters(sigma-70), the complete result

hasn’t been analyzed yet, however online promoter finders didn’t back up the hypothesis of the sequence being a promoter. Phage promoters don’t have a complete database so the result is expected.

The group has written an abstract for the project during lab after discussion:

Repeats in genomics have a wide variety of significance, from signifying the start of a transposable element to being the terminator for a gene. In a newly sequenced and annotated bacteriophage, NapoleonB, a 46-base pair sequence that was repeated twice was found at around 25,500bp and 26,165bp in a non-coding region. The sequence was analyzed to see if evidence was present for multiple functional elements, as well as compared to other AM cluster phages to search for similarity. Tools such as Gepard, NCBI’s BLAST, Clustal Omega, IS Finder, attP site finders, and others were used to try and match the sequence of interest to a potential function. The sequence was also found to have 4-6 base pair flanking inverted repeats, which piqued research interest towards a possible transposable element or promoter. Using the previously mentioned tools, we were able to rule out the notion that the sequence was a transposon, attP site, or an overlooked ORF or protein. However, with the sequence found to have been conserved across 11 of 14 AM cluster phages, it was concluded that the likelihood of the sequence occurring as observed at random was extremely low. Therefore, the leading remaining hypothesis that this repeat could function as a regulatory sequence or promoter seemed promising. In order to definitively confirm or rule out the possibility of a regulator sequence at this stage, the sequence would have to be removed in an in vitro setting and the following genes observed.

The Next Step:

The next lab would be to analyze the data in DNA Master and hopefully find supporting evidence for the hypothesis.