Parent Prochlorococcus NATL2A  transfer 0 replicate 1, PacBio WGS
Culture samples were prepared by pelleting 90 mL of exponentially growing cultures by centrifugation at 7,500 xg for 15 minutes. The pellet was resuspended into 200 l of pH 8 TE and stored at -80C. DNA was extracted using the same method as previously described above. Library construction began with diluting 4g of intact genomic DNA in 150 l of TE buffer (10 mM TrisHCl, 1 mM EDTA, pH 8.5) and fragmenting to 10-12 kb lengths using a gTube (Covaris Cat# 520079). Fragmented DNA was concentrated by a 0.45X SPRI bead cleanup (Beckman-Coulter), eluted in 37 l EB (10 mM TrisHCl pH 8.0) and processed using SMRTBellTM Template Prep kit 1.0 (PacBio Part# 100-259-100) to SMRTbell libraries, following manufacturer guidelines. Indexed blunt-end SMRTbell adapters (PacBio Part# 100-4666-000) were added for multiplexing. Multiple indexed SMRTbell libraries were assembled in a single pool, cleaned up using a Minelute reaction cleanup kit (QIAGEN Cat# 28204) to 100 l volume in EB, followed by SPRI 0.4X cleanup, with final elution in 11 l EB. Concentration and quality were assessed by Picogreen (ThermoFisher QuantIt Cat# P11496) and FEMTOpulse (Agilent). Pooled libraries were bound using Sequel Binding Kit 2.1 and sequenced for 10 h on the Pacific Biosciences Sequel I using a SMRTCell 1Mv2.