Hawaii Gigas Methylation Analysis Part 8
WGS resequencing quote
I plan on extracting SNP data from my WGBS data, but Steven suggested I get a quote for WGS resequencing. This would give us more robust genotype data that we could use in our methylation analysis. While there is the possibility Sam or I could extract additional DNA, I first asked Sam if there was any leftover ctenidia DNA. He said there was and shared this lab notebook post with the yield for each sample. According to this other post, he used 1500 µg of DNA for each sample. I quickly calculated how much DNA is left for each sample.
Table 1. DNA left for each ctenidia sample
|Sample_ID||Concentration(ng/uL)||Volume(uL)||Total_DNA (ng)||Amount Left (ng)|
Most samples have > 1000 µg of DNA, but there are three (2N_HI_11, 2N_LOW_5, 2N_LOW_6) that have less. Two samples (2N_HI_8 and 2N_HI_12) do not have any DNA left over, and they are both from the same treatment, so that could pose an issue. It’s interesting that the samples with less DNA are all diploid!
In any case, I submitted quotes to GENEWIZ and Northwest Genomics Center (NWGC). I asked how much DNA is required to for library preparation and sequencing. I spoke with Katie about WGS, and she suggested I base my quote on information from the Illumina coverage calculator. I’ll require need ~22 billion bases of output for the NovaSeq. I don’t know if GENEWIZ or NWGC use NovaSeq, but Katie thinks that sequencing will cost ~$6,000 for all samples.
I’ll update this issue with information when I get it.
- Create covariate matrix and complete pairwise DML assessment in
- Try BS-SNPer and EpiDiverse for SNP extraction from WGBS data
- Investigate comparison mechanisms for samples with different ploidy in oysters and other taxa
- Test-run DSS and ramwas
- Transfer scripts used to a nextflow workflow
- Update methods
- Update results