PacBio sequence reads of the Malassezia pachydermatis 23-36847-71 (LMC_L1) genome
M. pachydermatis strain 23-36487-71 (LMC_L1) was grown to saturation at 35C in liquid modified Dixon medium (ATCC Medium 2693). Cells were collected by centrifugation, washed in sterile water and resuspended in 1.5 ml of Solution 3 and 1.5 ml of Solution 4 from the genomic DNA prep described at dx.doi.org/10.17504/protocols.io.rm7vzb1b4vx1/v1. Three ml of phenol/chloroform (1:1) and a 0.5 ml equivalent of sterile glass beads (425-600 microns) were added to the tube. The tube was vortexed at top speed for 2 min, then centrifuged at 3000 x g to separate the phases. The aqueous phase was collected and DNA precipitated with 2 volumes of isopropanol. DNA was resuspended in 10 mM Tris-EDTA (pH 8.0), then treated with RNase A (Gold Biotechnology) at 37C for 1 h. Proteinase K (Gold Biotechnology) was added and incubated at 50C for 16 h. The preparation was extracted with phenol/chloroform and DNA precipitated with 2 volumes of isopropanol. Genomic DNA was sheared to a mean length of 15 kb using a Megaruptor 3 (Diagenode). Sheared DNA was run on a Blue Pippin system (Sage Science) to select 7-50 kb fragments using a 0.75% agarose cassette and DNA marker S1. Sheared fragments were converted to a library using the SMRTbell prep kit 3.0 (Pacific Biosciences) and SMRTbell adaptor index plate 96D. DNA sequencing was performed with SPARQ chemistry in a Revio instrument with a 30-h movie. CCS analysis was done on the instrument (ccs --min-passes 3 min-rq 0.99). Demultiplexing used SMRTlink v25.1.