Fig 1.
Workflow for extraction of HMW gDNA from Botryococcus.
Created with BioRender.com.
Fig 2.
Images of ground Botryococcus biomass in sorbitol wash buffer before and after centrifugation.
(A), (B), (C) Homogenized biomass resuspended in sorbitol wash buffer before centrifugation for each Botryococcus species. (D), (E), (F) Separation of biomass phases in sorbitol wash buffer after centrifugation. f, floating biomass phase; s, liquid supernatant; p, biomass pellet phase. The f and p phases are saved after each wash step.
Table 1.
Raw sequencing metrics obtained on the Oxford Nanopore PromethION platform from a single multiplexed flow cell run.
Number of Reads: total number of reads obtained; Total BP: total number of base pairs sequenced; Max Read Size: size of the longest read in bp; N50 BP: total length in bp that equals 50% of the Total BP; N50 NUM: number of reads used to obtain the N50 BP value; N90 BP: total length in bp that equals 90% of the Total BP; N90 NUM: number of reads used to obtain the N90 BP value; MEAN: mean read length, MEDIAN: median read length; Coverage: ratio of Total BP to genome size.
Fig 3.
Qualitative and quantitative analysis of HMW gDNA extracted from the three Botryococcus species.
(A) 0.5% agarose gel electrophoresis of 200 ng HMW gDNA from each Botryococcus species. (B) DNA concentration based on A260 analysis. (C) A260/230 ratio to assess polysaccharide contamination. (D) A260/280 ratio to assess protein contamination. In (C) and (D), data represents the mean ± standard error of four independent samples for B. alkenealis and B. lycopadienor, and six samples for B. braunii. A one-way ANOVA with the Friedman test was performed, and no significant differences were found between samples. (E) Size distribution of extracted HMW gDNA using an Agilent Femto Pulse system. Vertical dashed line indicates HMW gDNA ≥50kb in each sample.