De Novo Genome Assembly of Paraburkholderia kirstenboschensis

Genomic DNA was extracted from P. kirstenboschensis F3 that was grown in ISP2 for 18 h, using a modified phenol/chloroform extraction, where RNase A was added during the lysozyme incubation step [67 (link)]. DNA quality was confirmed using gel electrophoresis, Bioanalyzer, a NanoDrop One UV–Vis spectrophotometer, and a Qubit fluorometer (Invitrogen Qubit DNA-HS assay kit, Q32851). Genomic DNA was run through the UCB QB3 PacBio Sequel II sequencing pipeline. Raw reads were partitioned using seqtk, and the genome was assembled de novo with Flye into three contigs at 989x coverage. The draft genome was annotated using the NCBI Prokaryotic Genome Annotation Pipeline [68 (link)].
Species identification was determined by ANI analysis using the Kostas lab ANI calculator [69 ] in comparison to the NCBI reference genome for P. kirstenboschensis (GenBank accession number GCA_904848585.1). P. kirstenboschensis was among a list of Paraburkholderia species that had a > 97% identity match from BLAST searches using 16S rRNA, gyrB, recA, rpoB, and trpB DNA sequences from strain F3. Only P. kirstenboschensis had an ANI of >95%, an acceptable cutoff for bacterial species identification [70 (link)], with a resulting 96.83% ANI.