The effect of variant interference on de novo assembly for viral deep sequencing

Christina J. Castro; Rachel L. Marine; Edward Ramos; Terry Fei Fan Ng

The copyright holder for this preprint (which was not peer-reviewed) is the author/funder. Simulated genomes were generated using custom scripts that randomly assign each nucleotide over a 284 designated genome length with a weighted distribution dependent on the GC content [Supplement Figure S1 ]. 285 The random genomes were then screened using NCBI BLAST to insure no similarity/identity existed to any 286 classified organism (i.e., no BLAST hits). These simulated genomes served as the initial variant genome (variant 287 1). To generate the mutated variant genomes (variant 2), a custom script was used to systematically introduce 288 evenly distributed random mutations at rates from 1 mutation in every 4 nucleotides (75% PID) to 1 mutation 289 in every 250 nucleotides (99.6% PID), incrementing by 1 nucleotide. 290 291 Following the generation of initial and mutated variant genomes, high-quality fastq reads were 292 generated using ART, 33 simulating Illumina MiSeq paired-end runs at 50X coverage with 250 nt reads, 293 DNA/RNA mean fragments size of 500, and quality score of 93. Fastq reads were combined in equal numbers 294 for the initial and mutated variants, and used as input for subsequent de novo assembly experiments 295 [Supplement Figure S1 ]. The same process was utilized to generate the artificial genomes, initial and mutated 296 variant genomes, and reads for each of the experiments. 297 All rights reserved. No reuse allowed without permission.