Question: Task 9 5% The Next Step Is To Perform Genotypic Characterization. Here We Will Only Focus On The Unknown Bacterium. The First Step Is To Sequence The 16S RRNA Gene. Once Sequenced It Is Compared (or A BLAST) To 16S RRNA Sequences Available On A Database (NCBI Is The One Most Commonly Used). The Unknown Bacterium Has A 95% Similarlity To An Undescribed …

Task 9 5% The next step is to perform genotypic characterization. Here we will only focus on the unknown bacterium. The first step is to sequence the 16S rRNA gene. Once sequenced it is compared (or a BLAST) to 16S rRNA sequences available on a database (NCBI is the one most commonly used). The unknown bacterium has a 95% similarlity to an undescribed Proteus sp. (the cut-off value for a genus is <94%). We now need to identify the species. Housekeeping genes such as dnaj, mdh, pyro, recA and rpoD were sequenced, a concatenated (combined) phylogenetic tree (represents evolutionary relationships among organisms) was drawn (see next page). You will notice from the tree that the unknown bacterium groups separately to all the others thus indicating that it is a novel species. To confirm this observation, the genome of the isolate, which we will call MBY2021, needs to be sequenced, the C+G%, average nucleotide index (ANI) and web-based DNA:DNA hybridisation values determined. The C+G% was calculated at 56%. The dDDH and ANI values of MBY2021 was 80.8% and 23.5% , less than the proposed cutoff level for species delineation, i.e. 95 and 70%, respectively. Why is the 16S rRNA gene sequenced and not another gene? What are housekeeping genes? What is multilocus sequence analysis (MLSA)? What is a phylogenetic tree? What is dDDH and ANI? What is it necessary to determine these values ?