The PhD student's long careful reanalysis of the DNA uptake data has finally produced uptake ratio plots.  These confirm a surprising difference between the DNAs from two closely related strains, 86-028NP ('NP') and PittGG ('GG').  We also saw this difference in our preliminary analysis, but we thought it might be an artefact of how the analysis was done.In the experiment underlying this data, cells of a third strain, KW20, took up DNA that had been…

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This is a continuation of the previous post.For each of our 12 genuinely contaminated uptake samples we want to create multiple replicate fake-contaminated input samples, each fake-contaminated with an independent set of Rd reads at that sample's level of contamination.For example, our UP01 sample has 5.3% Rd contamination.  Its corresponding input sample is UP13.  UP13 has about 2.7 x10^6 reads, so to make a fake-contaminated sample for UP13 we need to add (2.7x10^6 * 0.053)/(1-0.053)…

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Yesterday the PhD student showed me the results of his contamination-correction tests.  They confirmed that our new error-correction strategy works, and suggested an improvement.The problem and the strategy:  We want to know how efficiently different segments of NP or GG DNA are taken up by competent Rd cells.  All of our 12 'uptake' samples are contaminated, consisting of mostly reads of NP or GG DNA taken up by Rd cells plus varying amounts of contaminating…

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Yesterday I went to an information session about UBC's Faculty Pension Plan, designed for faculty approaching the age when we have to decide what to do with the money the Plan has accumulated on our behalf.  Even if we choose not to retire (or not yet), by age 71 contributions to the Plan will end and we must make a decision.Until retirement or age 71, the Plan has been taking money from our salaries and…

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Yesterday I went to an information session about UBC's Faculty Pension Plan, designed for faculty approaching the age when we have to decide what to do with the money the Plan has accumulated on our behalf.  Even if we choose not to retire (or not yet), by age 71 contributions to the Plan will end and we must make a decision.Until retirement or age 71, the Plan has been taking money from our salaries and…

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In the previous post I proposed an alternative method to control for contaminating recipient DNA in our donor DNA uptake samples.  Because we've never done this before (and probably nobody else has either), we need ways to check that it accomplishes what we want.Here's one way: We already have made samples of pure donor DNA reads (from strain NP or GG) that have been deliberately contaminated with reads from the recipient Rd (10% Rd, 90%…

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Yesterday's post considered the factors affecting our current strategy for removing the contamination Rd-derived reads from our 'uptake' samples.  Since then I've looked at the read-mapping results using our new perfected reference sequences.  The problem of reads that have no SNPs and thus map ambiguously to both Rd and NP/GG is worse than we thought.  Below I'll describe this data and the new strategy I want to consider.The former post-doc sent us a spreadsheet with…

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Yesterday's post considered the factors affecting our current strategy for removing the contamination Rd-derived reads from our 'uptake' samples.  Since then I've looked at the read-mapping results using our new perfected reference sequences.  The problem of reads that have no SNPs and thus map ambiguously to both Rd and NP/GG is worse than we thought.  Below I'll describe this data and the new strategy I want to consider.The former post-doc sent us a spreadsheet with…

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We finally have the corrected reference genome sequences for our DNA uptake deep-sequencing data.  The genome sequences from NCBI had errors and polymorphisms relative to the specific strains we used in the experiments, so the former post-doc used the sequence data from this experiment to create reference sequences that perfectly match the DNAs we used.The bright-coloured table below shows 8 of our 16 samples.  These are the samples that examined uptake of DNA from strain…

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We finally have the corrected reference genome sequences for our DNA uptake deep-sequencing data.  The genome sequences from NCBI had errors and polymorphisms relative to the specific strains we used in the experiments, so the former post-doc used the sequence data from this experiment to create reference sequences that perfectly match the DNAs we used.The bright-coloured table below shows 8 of our 16 samples.  These are the samples that examined uptake of DNA from strain…

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