Whole genome sequencing of Mycoplasma pneumoniae strain 309
Mycoplasma pneumoniae strain 309 was cultured in 20 ml of PPLO medium at 37˚C for 5 days. M. pnenmoniae cells attached to the inner surface of the culture flask were scraped out and washed with phosphate buffered saline (PBS) twice. Cells were concentrated by centrifuge and suspended in 0.2 ml of PBS. Genomic DNA was extracted from cells using QIAamp DNA Mini Kit. Extracted DNA (26 ng/ micro liter) was diluted and sheared by Covaris S220 system according to manufacture’s instructions. Sheared DNA was purified by MinElute PCR Purification Kit and subjected to library preparation and 454 pyrosequencing according to manufacture’s instructions.