Daily Bits - February 2024

2024
Daily Bits
Author

Sam White

Published

February 1, 2024

20240229

Lab Meeting

  • Discussed sections from “Braiding Sweetgrass”

CEABIGR (GitHub repo)

  • Finalized females/males controls/exposed expression table.

  • Weekly meeting.

  • Created faceted volcano plots of female/male control/exposed transcript and gene expression:

Volcano plots of transcript/gene expression in female/male control/exposed samples. Black points are genes/transcripts with p/q-values > 0.05. Females controls transcripts are the only group to exhibit any differential expression, and can be seen as the single blue point, labelled with the gene ID LOC111134855.

Volcano plots of transcript/gene expression in female/male control/exposed samples. Black points are genes/transcripts with p/q-values > 0.05. Females controls transcripts are the only group to exhibit any differential expression, and can be seen as the single blue point, labelled with the gene ID LOC111134855.”

20240228

Miscellaneous

  • Read designated sections of “Braiding Sweetgrass” for lab meeting.

CEABIGR (GitHub repo)

  • Created supplemental table with female/male controls/exposed expression info.

20240227

Miscellaneous

CEABIGR (GitHub repo)

  • Generated table with mean FPKM values for genes in female/male control/exposed.

20240226

In lab

  • Selected and prepared primers for Eric’s qPCRs.

  • Helped Eric conduct qPCRs.

  • Helped Katherine gather sampling supplies for hatchery.

  • Ran all over the place in Bagley Hall to get LN2! Chemistry Stores was out and so was secondary location in Rm. 305A. Eventually managed to get LN2 in 405A…

CEABIGR (GitHub repo)

  • Generated canonical predominant isoform table.

20240223

Coral E5 (GitHub repo)

  • All hands meeting.

  • Read original proposal.

  • Read paper about plant epigenetic stress responses.

Science Hour

CEABIGR (GitHub repo)

  • Created canonical max transcripts per gene table for females controls/exposed and males controls/exposed.

20240222

Lab Meeting

  • Discussed Pub-a-thon 2024

CEABIGR (GitHub repo)

  • Reviewed recent GitHub Issues

  • Worked on creating canonical gene/transcript counts table.

Miscellaneous

  • Updated Nightingales (Google Sheet) spreadsheet with Cod RNA-seq info.

  • Raven was bogged down by some weird rogue git command, so had to reboot.

20240221

Miscellaneous

CEABIGR (GitHub repo)

  • Generated volcano plots

Volcano plot of gene expression in females. No genes are differentially expressed. Grey points are genes with p-values <=0.05. Black points are genes with p/q-values > 0.05.

Volcano plot of gene expression in females. No genes are differentially expressed. Grey points are genes with p-values <=0.05. Black points are genes with p/q-values > 0.05.

Volcano plot of transcript expression in females. A single transcript is differentially expressed and is rendered as a blue point and labelled with its associated geneID. Grey points are transcripts with p-values <=0.05. Black points are transcripts with p/q-values > 0.05.

Volcano plot of transcript expression in females. A single transcript is differentially expressed and is rendered as a blue point and labelled with its associated geneID. Grey points are transcripts with p-values <=0.05. Black points are transcripts with p/q-values > 0.05.

Volcano plot of gene expression in males. No genes are differentially expressed. Grey points are genes with p-values <=0.05. Black points are genes with p/q-values > 0.05.

Volcano plot of gene expression in males. No genes are differentially expressed. Grey points are genes with p-values <=0.05. Black points are genes with p/q-values > 0.05.

Volcano plot of transcript expression in males. No transcripts are differentially expressed. Grey points are transcripts with p-values <=0.05. Black points are transcripts with p/q-values > 0.05.

Volcano plot of transcript expression in males. No transcripts are differentially expressed. Grey points are transcripts with p-values <=0.05. Black points are transcripts with p/q-values > 0.05

20240220

CEABIGR (GitHub repo)

  • Started working on generating volcano plots.

20240216

Coral E5 (GitHub repo)

  • Finished running ShortStack variations with newly trimmed P.meandrina sRNA-seq.

20240215

Coral E5 (GitHub repo)

  • Ran ShortStack variations with newly trimmed P.meandrina sRNA-seq.

20240214

Coral E5 (GitHub repo)

20240213

Coral E5 (GitHub repo)

  • Cranked out new trimming on P.meandrina sRNA-seq samples.

20240212

In lab

  • Reviewed reverse transcription and qPCR with Eric.

Coral E5 (GitHub repo)

  • Started running new trimming on A.pulchra sRNA-seq samples (preivously, had just tried with P.evermanni).

20240209

Science Hour

  • Steven and I looked at and discussed the exon expression patterns of individual genes and compared those patterns to the WGCNA outputs which Ariana generated at yesterday’s meeting. Definitely a few individuals whose exon usage seems to deviate a great deal from that WGCNA module assignment…

Coral E5 (GitHub repo)

  • Continued to work on implementing new trimming/merging parameters, but ShortStack keeps encountering an error (GitHub Issue). I’m hoping the developer can lend some insight. This is weird, because these samples, albeit trimmed with a different program and used as paired-end reads (only trying R1 or merged reads at this point), were successfully analyzed previously…

20240208

Lab meeting

CEABIGR (GitHub repo)

  • Special, extended session with Arianna in attendance. She ran some WGCNA (PDF) analysis to try to see how exon expression patterns varied between control/exposed samples. Very cool!

20240207

Coral E5 (GitHub repo)

  • Finished messing around with trimming/merging stuff. Here’s the notebook entry.

  • Began implementing new trimming/merging.

20240206

Coral E5 (GitHub repo)

  • Continued to explore trimming and merging strategies for sRNA-seq data, using fastp alone, BBMerge alone, and/or combining the two approaches.

CEABIGR (GitHub repo)

  • Spent a few hours on a video chat with Steven and Yaamini going through exon fold-change code/analysis.

20240205

Coral E5 (GitHub repo)

  • Continued to explore trimming and merging strategies for sRNA-seq data, using fastp alone, BBMerge alone, and/or combining the two approaches.

20240202

Coral E5 (GitHub repo)

Science Hour

  • Steven and I played around with exon fold-change data.

20240201

In Lab

  • Lab meeting.

  • Helped Eric get started with RNA quantification using Qubit.

Coral E5 (GitHub repo)

  • Tested out more R1/R2 merging/trimming using fastp.

    • Merging is similar to BBMerge results - i.e. a (relatively) small number of reads are actually merging, despite the fact that these should be short insert sizes and R1/R2 reads should overlap…

  • Installed XICRA (Sanchez Herrero et al. 2021) on Raven to identify isomiRs in sRNA-seq data. Possibly to get around the merging “issues” experienced with BBMerge and fastp.

    • Installation was a bit bumpy, due to some weird Conda/Mamba “flexible solve” problem, but got that straightened out with conda config --set channel_priority flexible.

    • Installation test(s) had issues. I created the following GitHub Issues on the developer’s site in hopes of getting help:

    • installer.sh error

    • “No samples were retrieved” in test data

CEABIGR (GitHub repo)

  • Need to generate R2 values and slope of exon expression for all genes, Exons 1 - 6.

References

Sanchez Herrero, Jose Francisco, Raquel Pluvinet, Antonio Luna de Haro, and Lauro Sumoy. 2021. “Paired-End Small RNA Sequencing Reveals a Possible Overestimation in the isomiR Sequence Repertoire Previously Reported from Conventional Single Read Data Analysis.” BMC Bioinformatics 22 (1). https://doi.org/10.1186/s12859-021-04128-1.