phiM19_plaque_variant2 genome sequencing
A plaque variant of phage phiM19 with a 17,976 bp insertion after position 459 in the published phiM19 sequence. Our sequence of one plaque of phiM19 (phiM19.1) has 1 bp mismatch from the published phiM19 genome. The sequence of the second plaque of phiM19 (phiM19.2) is identical to the published phiM19 sequence from base 1-459, followed by a 17,976 bp insertion in phiM19.2 similar to the sequence of phiM12. 100% sequence identity between the published sequence of phiM19 and phiM19.2 resumes after the insertion. Two separate phiM19 DNA samples, each from a plaque-purified phage sample, were sheared with a Diagenode Bioruptor. Indexed libraries were constructed with an NEBNext Ultra DNA Library Prep kit for Illumina (NEB, Ipswich, MA) in accordance with the manufacturers instructions. For each library, 1 g of DNA was end repaired and ligated to NEBNext adapters. Fragments of an average size 860 bp for sample phiM19.2 and 1010 bp for sample phiM19.1 were isolated by electrophoresis on Bio-Rad Low Range Ultra Agarose and amplified for 8 to 10 cycles with NEB High-Fidelity 2X master mix and NEBNext multiplex oligonucleotides. Library size distribution was measured on an Agilent Bioanalyzer high-sensitivity chip, and quantity was determined with the KAPA Biosystems Library Quantification kit. Paired-end 300-base sequence reads were generated on an Illumina MiSeq with a 600-cycle MiSeq v3 Reagent kit. Genome assembly from MiSeq reads was performed with Lasergene SeqMan Pro v. 11.2.1.25 (DNAStar, Madison, WI). For both phiM19.2 and phiM19.1 the coverage was approximately 4000x. Both assemblies are circularly permuted.