Transcript Identification and Alignments - C.virginica RNAseq with NCBI Genome GCF_002022765.2 Using Hisat2 and Stringtie on Mox Again
2023
Miscellaneous
In the process of generating expression matrices for CEABIGR (GitHub repo), and in turn distance matrices for these samples, I realized a couple of things:
Reverse Transcription - C.gigas RNA from Noah’s Heat-Mechanical Stress Project
2023
Miscellaneous
After isolating ctenidia RNA on 20230712 and then quantifying the RNA earlier today, I needed to perform reverse transcription to have cDNA for subsequent qPCRs. Reverse transcription was performed using oligo dT primers using M-MLV RT (Promega), per the…
ORF Identification - L.staminea De Novo Transcriptome Assembly v1.0 Using Transdecoder on Mox
2023
Miscellaneous
After performing a de novo transcriptome assembly with L.staminea RNA-seq data, the
Trinity assembly stats were quite a bit more “exaggerated” than normally expected. In an attempt to get a better sense of which contigs might be…
FastQ QC and Trimming - E5 Coral RNA-seq Data for A.pulchra P.evermanni and P.meandrina Using FastQC fastp and MultiQC on Mox
2023
E5
After downloading and then reorganizing the E5 coral RNA-seq data from Azenta project 30-789513166, I ran FastQC for initial quality checks, followed by trimming with
fastp, and then final QC with FastQC/MultiQC. This was performed on all three species in the data…
Data Management - E5 Coral RNA-seq and sRNA-seq Reorganizing and Renaming
2023
Data Management
Downloaded the E5 coral sRNA-seq data from Azenta project 30-852430235 on 20230515 and the E5 coral RNA-seq data from Azenta project 30-789513166 on 20230516. The data required some reorganization, as the project included data from three…
Containers - Apptainer Explorations
2023
Miscellaneous
At some point, our HPC nodes on Mox will be retired. When that happens, we’ll likely purchase new nodes on the newest UW cluster, Klone. Additionally, the
coenv nodes are no longer available on Mox. One was decommissioned and one was “migrated” to Klone. The primary issue at hand is that the base…
Sequencing Read Taxonomic Classification - M.magister RNA-seq Using DIAMOND BLASTx and MEGAN6 daa-meganizer on Mox
2023
Miscellaneous
Running DIAMOND BLASTx, followed by MEGAN6
daa-meganizer for taxonomic classification of NOAA M.magister trimmed RNA-seq reads (provided by Giles Goetz on 20230301). This is primarily just for…
FastQ Trimming and QC - P.verrucosa RNA-seq Data from Danielle Becker in Hollie Putnam Lab Using fastp FastQC and MultiQC on Mox
2023
E5
After receiving the P.verrucosa RNA-seq data from Danielle Becker (Hollie Putnam’s Lab, Univ. of Rhode Island), I noticed that the trimmed reads didn’t appear to actually be trimmed. There was still adapter contamination…
Data Received - P.verrucosa RNA-seq and WGBS Full Data from Danielle Becker
2023
E5
Data Received
Worked with Danielle Becker, as part of the Coral E5 project, to transfer data related to her repo, from her HPC (Univ. of Rhode Island; Andromeda) to ours (Univ. of Washington; Mox) in order to eventually transfer to Gannet so…
Data Wrangling - P.acuta Genome GFF to GTF Conversion Using gffread
2023
Miscellaneous
As part of getting these three coral species genome files (GitHub Issue) added to our Lab Handbook Genomic Resources page, I will index the P.acuta genome file using
HISAT2, but need a GTF file to also identify exon/intro splice sites. Since a GTF file is not available, but a GFF file is, I needed to convert the GFF to GTF.… 
Data Wrangling - C.virginica NCBI GCF_002022765.2 GFF to Gene and Pseudogene Combined BED File
2022
Miscellaneous
Working on the CEABIGR project, I was preparing to make a gene expression file to use in CIRCOS (GitHub Issue) when I realized that the Ballgown gene expression file (CSV; GitHub) had more…
BSseq SNP Analysis - Nextflow EpiDiverse SNP Pipeline for C.virginica CEABIGR BSseq data
2022
Miscellaneous
Steven asked that I identify SNPs from our C.virginica CEABIGR BSseq data (GitHub Issue). So, I ran sorted, deduplicated Bismark BAMs that Steven generated through the EpiDiverse/snp Nextflow…
RNAseq Alignments - P.generosa Alignments and Alternative Transcript Identification Using Hisat2 and StringTie on Mox
2022
Miscellaneous
As part of identifying long non-coding RNA (lncRNA) in Pacific geoduck(GitHub Issue), one of the first things that I wanted to do was to gather all of our geoduck RNAseq data and align it to our geoduck genome. In addition to the alignments…
FastQ Trimming - Geoduck RNAseq Data Using fastp on Mox
2022
Miscellaneous
Per this GitHub Issue, Steven asked me to identify long non-coding RNA (lncRNA) in geoduck. The first step is to aggregate all of our Panopea generosa (Pacific geoduck) RNAseq data and get it all trimmed. After that, align it to the genome…
FastQ Trimming and QC - C.virginica Larval BS-seq Data from Lotterhos Lab and Part of CEABIGR Project Using fastp on Mox
2022
Miscellaneous
We had some old Crassostrea virginica (Eastern oyster) larval/zygote BS-seq data from the Lotterhos Lab that’s part of the CEABiGR Workshop/Project (GitHub Repo) and…
Splice Site Identification - S.namaycush Liver Parasitized and Non-Parasitized SRA RNAseq Using Hisat2-Stingtie with Genome GCF_016432855.1
2022
Miscellaneous
After previously downloading/trimming/QCing S.namaycush SRA liver RNAseq data on 20220706, Steven asked that I run through Hisat2 for splice site identification …
qPCR - Repeat of Mussel Gill Heat Stress cDNA with Ferritin Primers
2022
Miscellaneous
My previous qPCR on these cDNA using ferritin primers (SRIDs: 1808, 1809) resulted in no amplification. This was a bit surprising and makes me suspect that I screwed up somewhere (not adding primer(s)??), so I decided to…
Differential Gene Expression - P.generosa DGE Between Tissues Using Nextlow NF-Core RNAseq Pipeline on Mox
2022
Miscellaneous
Steven asked that I obtain relative expression values for various geoduck tissues (GitHub Issue). So, I decided to use this as an opportunity to try to use a Nextflow pipeline. There’s an RNAseq…
Data Analysis - C.virginica RNAseq Zymo ZR4059 Analyzed by ZymoResearch
2022
Miscellaneous
After realizing that the Crassostrea virginica (Eastern oyster) RNAseq data had relatively low alignment rates (see this notebook entry from 20220224 for a bit more background), I contacted ZymoResearch to see if they had any insight on what might be happening. I suspected rRNA contamination. ZymoResearch was kind enough…
Trimming - Additional 20bp from C.virginica Gonad RNAseq with fastp on Mox
2022
Miscellaneous
When I previously aligned trimmed RNAseq reads to the NCBI C.virginica genome (GCF_002022765.2) on 20210726, I specifically noted that alignment rates were consistently lower for males than females. However, I let that discrepancy distract me from a the larger issue: low…
Transcriptome Assembly - Genome-guided C.virginica Adult Gonad OA RNAseq Using Trinity on Mox
2022
Miscellaneous
As part of this project, Steven’s asked that I identify long, non-coding RNAs (lncRNAs) (GitHub Issue) in the Crassostrea virginica (Eastern oyster) adult OA gonad RNAseq data we have. The initial step for this is to assemble transcriptome. I generated…
RNAseq Alignment - C.virginica Adult OA Gonad Data to GCF_002022765.2 Genome Using HISAT2 on Mox
2022
Miscellaneous
As part of this project, Steven’s asked that I identify long, non-coding RNAs (lncRNAs) (GitHub Issue) in the Crassostrea virginica (Eastern oyster) adult OA gonad RNAseq data we have. The initial step for this is to assemble…
Data Wrangling - C.virginica Gonad RNAseq Transcript Counts Per Gene Per Sample Using Ballgown
2022
Miscellaneous
As we continue to work on the analysis of impacts of OA on Crassostrea virginica (Eastern oyster) gonads via DNA methylation and RNAseq (GitHub repo), we decided to compare the number of transcripts expressed per…
RNA Isolation - O.nerka Berdahl Tissues
2021
Miscellaneous
Finally got around to tackling this GitHub issue regarding isolating RNA from some Oncorhynchus nerka (sockeye salmon) tissues we have from Andrew Berdahl’s lab (a UW SAFS professor) to use for RNAseq and/or qPCR. We have blood, brain, gonad, and liver samples from individual salmon from two different groups: territorial and social…
SNP Characterization - C.bairdi v3.1 Transcriptome Assembly and Day2 DEG Pooled Samples RNAseq Data
2021
Tanner Crab RNAseq
I previously identified variants across the four Day 2 DEG pooled RNAseq samples (380822, 380823, 380824, 380825) on 20210909 within the
cbai_transcriptome_v3.1 transcriptome assembly. Now, I needed to actually do some analysis on the SNPs for the…
RNAseq Alignments - C.bairdi Day 2 Infected-Uninfected Temperature Increase-Decrease RNAseq to cbai_transcriptome_v3.1.fasta with Hisat2 on Mox
2021
Tanner Crab RNAseq
Ealier today, I created the necessary Hisat2 index files for
cbai_transcriptome_v3.1. Next, I needed to actually get the alignments run. The alignments were performed using HIS…
Assembly Indexing - C.bairdi Transcriptome cbai_transcriptome_v3.1.fasta with Hisat2 on Mox
2021
Tanner Crab RNAseq
…
Transcript Identification and Quantification - C.virginia RNAseq With NCBI Genome GCF_002022765.2 Using StringTie on Mox
2021
Miscellaneous
After having run
HISAT2 to index and identify exons and splice sites in the NCBI Crassostrea virginica (Eastern oyster) genome (GCF_002022765.2) on 20210720, the next step was to identify and quantify transcripts from… 
Genome Assembly - Olurida_v090 with BGI Illumina and PacBio Hybrid Using Wengan on Mox
2021
Olympia Oyster Genome Assembly
I was recently tasked with adding annotations for our Ostrea lurida genome assembly to NCBI. As it turns out, adding just annotation files can’t be done since the genome was initially…
Transcriptome Assessment - BUSCO Metazoa on C.bairdi Transcriptome v4.0 on Mox
2021
Tanner Crab RNAseq
I previously created a C.bairdi de novo transcriptome assembly v4.0 with Trinity from all our C.bairdi RNAseq reads which had BLASTx matches to the C.opilio genome and decided to assess its “completeness” using…
DIAMOND BLASTx - C.bairdi RNAseq vs C.opilio Genome Proteins on Mox
2021
Tanner Crab RNAseq
We want to generate an additional Tanner crab (Chionoecetes bairdi) transcriptome, per this GitHub issue, to generate an additional C.bairdi transcriptome. This has come about due to the release of the genome of a very closely…
Data Wrangling - Gene ID Extraction from P.generosa Genome GFF Using Methylation Machinery Gene IDs
2021
Miscellaneous
Per this GitHub issue, Steven provided a list of methylation-related gene names and wanted to extract the corresponding Panopea generosa ([Pacific geoduck (Panopea generosa)](http://en.wikipedia.org/wiki/Geodu…
Data Wrangling - Gene ID Extraction from P.generosa Genome GFF Using Methylation Machinery List
2021
Miscellaneous
Per this GitHub Issue Steven asked that I take a list of gene names associated with DNA methylation and see if I could extract a list of Panopea generosa (Panopea generosa) gene IDs and corresponding BLAST e-values for each from…
Data Received - Anthopleura elegantissima - aggregating anenome - NanoPore Genome Sequence from Jay Dimond
2021
Miscellaneous
Jay asked me to help get his A.elegantissima (aggregating anenome) NanoPore gDNA sequencing data submitted to NCBI Sequencing Read Archive (SRA). He sent a hard drive (HDD) with all the NanoPore sequencing Fast5 files. The HDD was received on 2/2/2021. Here’re are details provided in the…
Alignments - C.bairdi RNAseq Transcriptome Alignments Using Bowtie2 on Mox
2020
Tanner Crab RNAseq
I had previously attempted to compare all of our C.bairdi transcriptome assemblies using DETONATE on 20200601, but, due to hitting time limits on Mox, failed to successfully get the analysis to complete. I realized that the limiting factor was performing FastQ alignments, so I…
Trimming - Haws Lab C.gigas Ploidy pH WGBS 10bp 5 and 3 Prime Ends Using fastp and MultiQC on Mox
2020
Miscellaneous
Making the assumption that the 24 C.gigas ploidy pH WGBS data we receved 20201205 will be analyzed using
Bismark, I decided to go ahead and trim the files according to Bismark guidelines…
Data Received - C.gigas Diploid-Triploid pH Treatments Ctenidia WGBS from ZymoResearch
2020
Data Received
Today we received the whole genome bisulfite sequencing (WGBS) from the 24 C.gigas diploid-triploid samples subjected to different pH that were submitted 20200824. The lengthy turnaround time was due to a bad lot of reagents, which forced them Zymo to find a different manufacturer in order to…
Sample Submission - M.magister MBD BSseq Libraries for MiSeq at NOAA
2020
Samples Submitted
Earlier today I quantified the libraries with the Qubit in preparation for sample pooling and sequencing. Before performing a full sequencing run, Mac wanted to select a subset of the libraries based on the experimental…
MBD BSseq Library Prep - M.magister MBD-selected DNA Using Pico Methyl-Seq Kit
2020
Miscellaneous
After finishing the final set of eight MBD selections on 20201103, I’m finally ready to make the BSseq libraries using the Pico Methyl-Seq Library Prep Kit (ZymoResearch) (PDF). I followed the manufacturer’s protocols with the following notes/changes (organized by each section in the…
Transcriptome Assessment - Crustacean Transcripome Completeness Evaluation Using BUSCO on Mox
2020
Miscellaneous
Grace was recently working on writing up a manuscript which did a basic comparison of our C.bairdi transcriptome (
cbai_transcriptome_v3.1) (see the Genomic Resources wiki for more deets) to two other species’ transcriptome assemblies. We wanted BUSCO evaluations as part of this comparison, but the two other species did not have BUSCO scores…
RNAseq Alignments - S.salar HISAT2 BAMs to GCF_000233375.1_ICSASG_v2_genomic.gtf Transcriptome Using StringTie on Mox
2020
Miscellaneous
This is a continuation of addressing Shelly Trigg’s (regarding some Salmo salar RNAseq data) request (GitHub Issue) to trim (completed 20201029), perform genome alignment (completed on…
MBD Selection - M.magister Sheared Gill gDNA 8 of 24 Samples Set 2 of 3
2020
Miscellaneous
Click here for notebook on the first eight samples processed. M.magister (Dungeness crab) gill gDNA provided by Mackenzie Gavery was previously sheared on 20201026 and three samples were subjected to additional rounds…
DNA Shearing - M.magister gDNA Shearing All Samples and Bioanalyzer
2020
Miscellaneous
I previously ran some shearing tests on 20201022 to determine how many cycles to run on the sonicator (Bioruptor 300; Diagenode) to achieve an average fragment length of ~350 - 500bp in preparation for MBD-BSseq. The…
DNA Shearing - M.magister CH05-21 gDNA Full Shearing Test and Bioanalyzer
2020
Miscellaneous
Yesterday, I did some shearing of Metacarcinus magister gill gDNA on a test sample (CH05-21) to determine how many cycles to run on the sonicator (Bioruptor 300; Diagenode) to achieve an average fragment length of ~350 - 500bp in…
DNA Shearing - M.magister gDNA Shear Testing and Bioanalyzer
2020
Miscellaneous
Steven assigned me to do some MBD-BSseq library prep (GitHub Issue) for some Dungeness crab (Metacarcinus magister) DNA samples provided by Mackenzie Gavery. The DNA was isolated from juvenile (J6/J7 developmental stages) gill tissue. One of the first steps in MBD-BSseq is to fragment…
Read Mapping - C.bairdi 201002558-2729-Q7 and 6129-403-26-Q7 Taxa-Specific NanoPore Reads to cbai_genome_v1.01.fasta Using Minimap2 on Mox
2020
Miscellaneous
After extracting FastQ reads using
seqtk on 20201013 from the various taxa I had been interested in, the next thing needed doing was mapping reads to the cbai_genome_v1.01 “genome” assembly from…
Taxonomic Assignments - C.bairdi 6129-403-26-Q7 NanoPore Reads Using DIAMOND BLASTx on Mox and MEGAN6 daa2rma on emu
2020
Miscellaneous
After noticing that the initial MEGAN6 taxonomic assignments for our combined C.bairdi NanoPore data from 20200917 revealed a high number of bases assigned to E.canceri and Aquifex…
Taxonomic Assignments - C.bairdi 20102558-2729-Q7 NanoPore Reads Using DIAMOND BLASTx on Mox and MEGAN6 daa2rma on emu
2020
Miscellaneous
After noticing that the initial MEGAN6 taxonomic assignments for our combined C.bairdi NanoPore data from 20200917 revealed a high number of bases assigned to E.canceri and Aquifex…
qPCR - Geoduck Normalizing Gene Primers 28s-v4 and EF1a-v1 Tests
2020
Miscellaneous
On Monday (20200914), I checked a set of 28s and EF1a primer sets and determined that 28s-v4 and EF1a-v1 were probably the best of the bunch, although they all looked great. So, I needed to test these out on some individual cDNA samples…
Samples Submission - Supplemental Ronits C.gigas Diploid-Triploid Ctendidia gDNA for WGBS by ZymoResearch
2020
Sample Submission
I re-quantified samples for Zymo whole genome bisulfite sequencing (WGBS) project 3534 earlier today due to ZymoResearch indicating there was insufficient DNA for most of the samples and submitted an additional 1ug of each…
DNA Quantification - Re-quant Ronits C.gigas Diploid-Triploid Ctenidia gDNA Submitted to ZymoResearch
2020
Miscellaneous
I received notice from ZymoResearch yesterday afternoon that the DNA we sent on 20200820 for this project (Quote 3534) had insufficient DNA for sequencing for most of the samples. This was, honestly, shocking. I had even submitted well over the minimum amount of DNA required (submitted…
Primer Design and In-Silico Testing - Geoduck Reproduction Primers
2020
Miscellaneous
Shelly asked that I re-run the primer design pipeline that Kaitlyn had previously run to design a set of reproduction-related qPCR primers. Unfortunately, Kaitlyn’s Jupyter Notebook wasn’t backed up and she accidentally…
SRA Submission - Geoduck Epigenetic Ocean Acidification RNAseq
2020
SRA Submission
Can’t remember where it was discussed (probably lab meeting), but I created a GitHub Issue to add all of geoduck RNAseq data to NCBI Short Read Archive (SRA). Anyway, got all the remaining RNAseq data uploaded to the NCBI SRA and organized into the correct…
ENA Submission - Ostrea lurida draft genome Olurida_v081.fa
2020
Olympia Oyster Genome Sequencing
Submitted our Ostrea lurida v081 genome assembly FastA to the European Nucloetide Archive.
Transcriptome Annotation - Trinotate C.bairdi Transcriptome-v1.7 on Mox
2020
Miscellaneous
After creating a de novo assembly of C.bairdi transcriptome v1.7 on 20200527, performing BLASTx annotation on 202000527, and TransDecoder for ORF identification on 20200527, I continued the annotation process by…
Transcriptome Comparisons - C.bairdi BUSCO Scores
2020
Miscellaneous
Since we’ve generated a number of versions of the C.bairdi transcriptome, we’ve decided to compare them using various metrics. Here, I’ve compared the BUSCO scores generated for each transcriptome using BUSCO’s built-in plotting script. The…
Transcriptome Assembly - C.bairdi All Pooled Arthropoda-only RNAseq Data with Trinity on Mox
2020
Miscellaneous
For completeness sake, I wanted to create an additional C.bairdi transcriptome assembly that consisted of Arthropoda only sequences from just pooled RNAseq data (since I recently generated a similar…
SRA Library Assessment - Determine RNAseq Library Strandedness from P.trituberculatus SRA BioProject PRJNA597187
2020
Miscellaneous
We’ve produced a number of C.bairid transcriptomes utilizing different assembly approaches (e.g. Arthropoda reads only, stranded libraries only, mixed strandedness libraries…
Transcriptome Annotation - Trinotate C.bairdi Transcriptome-v1.6 on Mox
2020
Miscellaneous
After creating a de novo assembly of C.bairdi transcriptome v1.6 on 20200518, performing BLASTx annotation on 202000519, and TransDecoder for ORF identification on 20200519, I continued the annotation process by…
Transcriptome Annotation - Trinotate C.bairdi Transcriptome v2.0 from 20200502 on Mox
2020
Miscellaneous
After performing de novo assembly on all of our Tanner crab RNAseq data (no taxonomic filter applied, either) on 20200502 and performing BLASTx annotation on 20200508, I continued the annotation process…
GO to GOslim - C.bairdi Enriched GO Terms from 20200422 DEGs
2020
Miscellaneous
After running pairwise comparisons and identify differentially expressed genes (DEGs) on 20200422 and finding enriched gene ontology terms, I decided to map the GO terms to Biological Process GOslims. Additionally, I decided to try another level of comparison (I’m not…
Transcript Abundance - C.bairdi Alignment-free with Salmon on Mox for Grace
2020
Miscellaneous
Per this GitHub Issue, Grace and Steven asked if I could help by generating a transcript abundance file for Grace to use with EdgeR. To do so, I used Salmon for alignment-free transcript abundance estimates due to its speed and…
Data Received - C.bairdi RNAseq Data from Genewiz
2020
Data Received
Tanner Crab RNAseq
We received the RNAseq data from the RNA that was sent out by Grace on 20200212.
NanoPore Sequencing - C.bairdi gDNA 6129_403_26
2020
Miscellaneous
After getting high quality gDNA from Hematodinium-infected C.bairdi hemolymph on 2020210 we decided to run some of the sample on the NanoPore MinION, since the flowcells have a very short shelf life. Additionally, the results from this will also help inform us…
qPCR - C.bairdi RNA Check for Residual gDNA
2020
Tanner Crab RNAseq
Previuosly checked existing crab RNA for residual gDNA on 20200226 and identified samples with yields that were likely too low, as well as samples with residual gDNA. For those samples, was faster/easier to just isolate more RNA and…
Trimming/MultiQC - Methcompare Bisulfite FastQs with fastp on Mox
2020
Miscellaneous
Steven asked me to trim a set of FastQ files, provided by Hollie Putnam, in preparation for methylation analysis using Bismark. The analysis is part of a coral project comparing DNA methylation profiles of different species, as well as comparing different sample…
RNA Isolation and Quantification - C.bairdi RNA from Hemolymph Pellets in RNAlater
2020
Tanner Crab RNAseq
Based on qPCR results testing for residual gDNA from 20200225, a set of 24 samples were identified that required DNase treatment and/or additional RNA. I opted to just isolate more RNA from all samples, since the kit includes a…
DNA Isolation & Quantification - Additional C.bairdi gDNA from Sample 6129_403_26
2020
Miscellaneous
Earlier today I isolated gDNA from C.bairi 6129_403_26 hemolymph pellets and recovered decently intact gDNA that could be used for sequencing. However, I still need more gDNA, so will isolate that (and co-isolate RNA, since I’m going through the…
Gene Expression - Hematodinium MEGAN6 with Trinity and EdgeR
2020
Miscellaneous
After completing annotation of the Hematodinium MEGAN6 taxonomic-specific Trinity assembly using Trinotate on 20200126, I performed differential gene expression analysis and gene ontology (GO) term enrichment analysis using Trinity’s scripts to run EdgeR and GOseq, respectively. The…
Gene Expression - C.bairdi MEGAN6 with Trinity and EdgeR
2020
Tanner Crab RNAseq
After completing annotation of the C.bairdi MEGAN6 taxonomic-specific Trinity assembly using Trinotate on 20200126, I performed differential gene expression analysis and gene ontology (GO) term enrichment analysis using Trinity’s scripts to run EdgeR and GOseq, respectively, across all…
RNA Isolation and Quantification - C.bairdi Hemocyte Pellets in RNAlater Troubleshooting
2020
Tanner Crab RNAseq
After the failure to obtain RNA from any C.bairdi hemocytes pellets (out of 24 samples processed) on 20200117, I decided to isolate RNA from just a subset of that group to determine if I screwed something up last time or something. Also, I am testing two different preparations of…
DNA Quality Assessment - Agarose Gel for C.bairdi 20102558-2729 gDNA from 20200122
2020
Miscellaneous
Earlier today, I isolated gDNA from C.bairdi 20102558-2729 ethanol-preserved muscle tissue using the Quick DNA/RNA MicroPrep Plus Kit (ZymoResearch) and prepared the tissue in three different ways to…
Data Wrangling - Arthropoda and Alveolata Taxonomic RNAseq FastQ Extractions
2020
Tanner Crab RNAseq
After using MEGAN6 to extract Arthropoda and Alveolata reads from our RNAseq data on 20200114 (for reference, these include RNAseq data using a newly established “shorthand”: 2018, 2019), I realized that the FastA…
RNAseq Reads Extractions - C.bairdi Taxonomic Reads Extractions with MEGAN6 on swoose
2020
Tanner Crab RNAseq
I previously ran BLASTx and “meganized” the output DAA files on 20200103 (for reference, these include RNAseq data using a newly established “shorthand”: 2018, 2019) and now need to use MEGAN6 to bin the results into the proper…
NanoPore Sequencing - Initial NanoPore MinION Lambda Sequencing Test
2020
Miscellaneous
We recently acquired a NanoPore MinION sequencer, FLO-MIN106 flow cell and the Rapid Sequencing Kit (SQK-RAD004). The NanoPore website provides a pretty thorough an user-friendly walk-through of how to begin using the system for the first time. With that said, I believe the user needs…
Trimming/FastQC/MultiQC - C.bairdi RNAseq FastQ with fastp on Mox
2019
Tanner Crab RNAseq
Grace/Steven asked me to generate a de novo transcriptome assembly of our current C.bairdi RNAseq data in this GitHub issue. As part of that, I needed to quality trim the data first. Although I could automate this as part of the transcriptome assembly (Trinity has Trimmomatic built-in), I would…
Samples Received - C.bairdi Hemolymph and Tissue in Ethanol from Pam Jensen
2019
Samples Received
Pam…
Data Wrangling - Rename Pgenerosa_v074 Bismark Coverage Files Scaffold Names
2019
Geoduck Genome Sequencing
After a meeting today, Steven and Hollie realized that the Bismark coverage files were still using the old scaffold names, stemming from the Pgenerosa_v074 naming. I’d previously updated filenames and…
Genome Feature Counts - Panopea-generosa-vv0.74.a4
2019
Geoduck Genome Sequencing
Due to a bug with GenSAS, the final GFF generated by their system to did not properly merge all of the repeats data properly. The repeats data below should be ignored! It appears that the other features data (e.g. CDS, exons, etc) was unaffected…
Data Received - C.bairdi RNAseq Day9-12-26 Infected-Uninfected
2019
Tanner Crab RNAseq
Data Received
Previously, we “received” this data, but it turns out it was incomplete (see 20191003)./
Genome Comparison - Pgenerosa_v074 vs Pgenerosa_v070 with MUMmer Promer on Mox
2019
Miscellaneous
In continuing to further improve our geoduck genome annotation, I’m attempting to figure out why Scaffold 1 of our assembly doesn’t have any annotations. As part of that I’ve decided to perform a series of genome comparisons and see how they match up, with an…
Genome Comparison - Pgenerosa_v074 vs S.glomerata NCBI with MUMmer Promer on Mox
2019
Miscellaneous
In continuing to further improve our geoduck genome annotation, I’m attempting to figure out why Scaffold 1 of our assembly doesn’t have any annotations. As part of that I’ve decided to perform a series of genome comparisons and see how they match up, with an…
Genome Comparison - Pgenerosa_v074 vs M.yessoensis NCBI with MUMmer Promer on Mox
2019
Miscellaneous
In continuing to further improve our geoduck genome annotation, I’m attempting to figure out why Scaffold 1 of our assembly doesn’t have any annotations. As part of that I’ve decided to perform a series of genome comparisons and see how they match up, with an…
Genome Comparison - Pgenerosa_v074 vs H.sapiens NCBI with MUMmer Promer on Mox
2019
Miscellaneous
In continuing to further improve our geoduck genome annotation, I’m attempting to figure out why Scaffold 1 of our assembly doesn’t have any annotations. As part of that I’ve decided to perform a series of genome comparisons and see how they match up, with an…
Genome Comparison - Pgenerosa_v074 vs C.virginica NCBI with MUMmer Promer on Mox
2019
Miscellaneous
In continuing to further improve our geoduck genome annotation, I’m attempting to figure out why Scaffold 1 of our assembly doesn’t have any annotations. As part of that I’ve decided to perform a series of genome comparisons and see how they match up, with an…
Genome Comparison - Pgenerosa_v074 vs C.gigas NCBI with MUMmer Promer on Mox
2019
Miscellaneous
In continuing to further improve our geoduck genome annotation, I’m attempting to figure out why Scaffold 1 of our assembly doesn’t have any annotations. As part of that I’ve decided to perform a series of genome comparisons and see how they match up, with an…
Genome Comparison - Pgenerosa_v074 vs Pgenerosa_v070 with MUMmer on Mox
2019
Miscellaneous
In continuing to further improve our geoduck genome annotation, I’m attempting to figure out why Scaffold 1 of our assembly doesn’t have any annotations. As part of that I’ve decided to perform a series of genome comparisons and see how they match up, with an…
Genome Comparison - Pgenerosa_v074 vs Pgenerosa_v074 with MUMmer on Mox
2019
Miscellaneous
In continuing to further improve our geoduck genome annotation, I’m attempting to figure out why Scaffold 1 of our assembly doesn’t have any annotations. As part of that I’ve decided to perform a series of genome comparisons and see how they match up, with an…
Genome Comparison - Pgenerosa_v074 vs S.glomerata NCBI with MUMmer on Mox
2019
Miscellaneous
In continuing to further improve our geoduck genome annotation, I’m attempting to figure out why Scaffold 1 of our assembly doesn’t have any annotations. As part of that I’ve decided to perform a series of genome comparisons and see how they match up, with an…
Genome Comparison - Pgenerosa_v074 vs M.yessoensis NCBI with MUMmer on Mox
2019
Miscellaneous
In continuing to further improve our geoduck genome annotation, I’m attempting to figure out why Scaffold 1 of our assembly doesn’t have any annotations. As part of that I’ve decided to perform a series of genome comparisons and see how they match up, with an…
Genome Comparison - Pgenerosa_v074 vs C.gigas NCBI with MUMmer on Mox
2019
Miscellaneous
In continuing to further improve our geoduck genome annotation, I’m attempting to figure out why Scaffold 1 of our assembly doesn’t have any annotations. As part of that I’ve decided to perform a series of genome comparisons and see how they match up, with an…
Genome Comparison - Pgenerosa_v074 vs H.sapiens NCBI with MUMmer on Mox
2019
Miscellaneous
In continuing to further improve our geoduck genome annotation, I’m attempting to figure out why Scaffold 1 of our assembly doesn’t have any annotations. As part of that I’ve decided to perform a series of genome comparisons and see how they match up, with an…
Genome Comparison - Pgenerosa_v074 vs C.virginica NCBI with MUMmer on Mox
2019
Miscellaneous
In continuing to further improve our geoduck genome annotation, I’m attempting to figure out why Scaffold 1 of our assembly doesn’t have any annotations. As part of that I’ve decided to perform a series of genome comparisons and see how they match up, with an…
RepeatMasker - Pgenerosa_v070 for Transposable Element ID on Roadrunner
2019
Geoduck Genome Sequencing
Continuing our various attempts at annotating our geoduck genome assemblies, I will be re-annotating our Pgenerosa_v070 (see Genome Resources GitHub wiki for deets) and realized I hadn’t run RepeatMasker on this assembly previously. Running RepeatMasker will generate a GFF that I can supply to MAKER to aid in repeats…
Genome Annotation - Pgenerosa_v074 Transcript Isoform ID with Stringtie on Mox
2019
Geoduck Genome Sequencing
After annotating Pgenerosa_v070 and comparing feature counts, there was a drastic difference between the two genome versions. Additionally, both of those genomes ended up with no CDS/exon/gene/mRNA features identified in the largest…
Genome Annotation - Pgenerosa_v070 Transcript Isoform ID with Stringtie on Mox
2019
Geoduck Genome Sequencing
After annotating Pgenerosa_v074 and comparing feature counts, there was a drastic difference between the two genome versions. Additionally, both of those genomes ended up with no CDS/exon/gene/mRNA features identified in the largest…
Genome Annotation - Pgenerosa_v070 Hisat2 Transcript Isoform Index
2019
Geoduck Genome Sequencing
This is the first step in getting transcript isoform…
Genome Annotation - Pgenerosa_v070 and v074 Top 18 Scaffolds Feature Count Comparisons
2019
Geoduck Genome Sequencing
After annotating Pgenerosa_v074 on 20190701, we noticed a large discrepancy in the number of transcripts that MAKER identified, compared to Pgenerosa_v070. As a reminder, the Pgenerosa_v074 is a subset of…
Genome Annotation - O.lurida 20190709-v081 Transcript Isoform ID with Stringtie on Mox
2019
Olympia Oyster Genome Sequencing
Earlier today, I generated the necessary Hista2 index, which incorporated splice sites and exons, for use with Stringtie in order to identify transcript isoforms in our 20190709-Olurida_v081…
Genome Assessment - BUSCO Metazoa on Pgenerosa_v074 on Mox
2019
Geoduck Genome Sequencing
Ran BUSCO on Mox for our Pgenerosa_v74 genome assembly to assess…
Genome Annotation - Olurida_v081 with MAKER and Tissue-specific Transcriptomes on Mox
2019
Olympia Oyster Genome Sequencing
I previously annotated our Olurida_v081 genome with MAKER using our “canonical” transcriptome, Olurida_transcriptome_v3.fasta as the EST evidence utilized by MAKER. A discussion on one of our Slack channels related to the lack of isoform…
Genome Annotation - O.lurida (v081) Transcript Isoform ID with Stringtie on Mox
2019
Olympia Oyster Genome Sequencing
Earlier today, I generated the necessary Hista2 index, which incorporated splice sites and exons, for use with Stringtie in order to identify transcript isoforms in our Olurida_v081…
RNA Isolation and Quantification - C.bairdi Hemolymph Pellet in RNAlater
2019
Miscellaneous
After the success we had isolating RNA using the Quick-DNA/RNA Microprep Plus Kit (ZymoResearch), Steven had me isolate RNA from a list of ~117 samples. Of that list, I was able to find 66 crab hemolymph pelleted RNAlater samples. The “missing” samples were most likely…
Data Analysis - C.virginica MBD Sequencing Coverage
2019
Miscellaneous
A while ago, Steven tasked me with assessing some questions related to the sequencing coverage we get doing MBD-BSseq in this GitHub issue. At the heart of it all was really to try to get an idea of how much usable data we actually get when we do an MBD-BSseq project. Yaamini happened to have done an MBD-BSseq…
Metagenome Assemblies - P.generosa Water Samples Trimmed HiSeqX Data Using Megahit on Mox to Compare pH Treatments
2019
Miscellaneous
A report involving our work on the geoduck water metagenomics is due later this week and our in-depth analysis for this project using Anvi’o is likely to require at least another…
RNA Isolation and Quantification - Crab Hemolypmh Using Quick-DNA-RNA Microprep Plus Kit
2019
Tanner Crab RNAseq
In the continuing struggle to isolate RNA from the Chionoecetes bairdi hemolymph preserved in RNAlater, Pam managed to find the Quick-DNA-RNA Microprep Plus Kit (ZymoResearch) as a potential option. We received a free sample of the kit and so…
Transcriptome Assembly - Geoduck Tissue-specific Assembly Larvae Day5 EPI99 with HiSeq and NovaSeq Data on Mox
2019
Miscellaneous
I previously assembled and annotated P.generosa larval Day 5 transcriptome (20190318 - mislabeled as Juvenile Day 5 in my previous notebook entries) using just our HiSeq data from our Illumina collaboration. This was a an…
Transcriptome Assembly - Geoduck Tissue-specific Assembly Juvenile Ambient OA EPI123 with HiSeq Data on Mox
2019
Miscellaneous
Ran a de novo assembly on our HiSeq data from Hollie’s juvenile EPI 123 sample “ambient OA”. This was done for Christian to use in some long, non-coding RNA (lncRNA) analysis.
Sample Submission - Lotterhos C.virginica Mantle MBD DNA to ZymoResearch for BSseq
2019
Miscellaneous
Crassostrea virginica samples that were subjected to MBD enrichment (completed 20190319) were shipped, on dry ice, to ZymoResearch today for BSseq. They will perform bisulfite conversion, library prep, and…
Data Management - SRA Submission Panopea generosa Day 5 Larvae RNAseq
2019
Miscellaneous
In preparation for some manuscripts, Steven asked that I get some geoduck RNAseq data upload to the NCBI sequencing read archive (SRA) in this GitHub Issue. Specifically, he needed the data corresponding to day 5 (pos-fertilization) larve RNAseq data that was prepped/run by Illumina on the NovaSeq. Original sample…
MBD Enrichment - C.virginica Sheared Mantle DNA from 20190306
2019
Miscellaneous
Last week I performed MBD enrichment on 12 of the 24 Lotterhos C.viriginica mantle DNA on samples I had sheared on 20190306. I proceeded with MBD enrichment using the MethylMiner Kit (Invitrogen) on the 12 remaining samples of the 24. I followed the manufacturer’s protocol for…
DNA Shearing & Bioanalyzer - Lotterhos C.virginica Mantle gDNA from 2018114
2019
Miscellaneous
Crassostrea virginica mantle gDNA that had previously been isolated on 20181114 was evaluated for integrity via agaroase gel yesterday (20190305). Everything looked pretty good, so proceeded with shearing in…
Genome Assessment - BUSCO Metazoa on P.generosa v071 on Mox
2019
Miscellaneous
Ran BUSCO on our completed annotation of the P.generosa v071 genome (GFF) (subset of sequences >10kbp). See this notebook entry for genome annotation info. This provides a nice metric on how “complete” a genome assembly (or transcriptome) is. Additionally…
Genome Annotation - Pgenerosa_v070 MAKER on Mox
2019
Geoduck Genome Sequencing
Here it goes, a massive undertaking of attempting to annotate the Pgenerosa_v070 genome (FastA; 2.1GB) using MAKER on Mox! I previously started this on 20190115, but killed it in order to fix a number of different issues with the script that were causing problems. I decided the changes were substantial enough, that I’d just make a…
DNA Isolation and Quantification - Ronit’s C.gigas Ploidy Ctenidia
2019
Miscellaneous
Last week, I isolated DNA from all of Ronti’s ctenidia samples, however one sample (D18-C) didn’t isolate properly. So, I performed another isolation procedure with another section of frozen tissue. Tissue was excised from frozen…
Annotation - Olurida_v081 MAKER ID Mapping
2019
Olympia Oyster Genome Sequencing
After getting through the initial MAKER annotation and SNAP gene predictions, I needed (wanted?) to simplify…
FastQC and Trimming - Metagenomics (Geoduck) HiSeqX Reads from 20180809
2018
Miscellaneous
Steven tasked me with assembling our geoduck metagenomics HiSeqX data. The first part of the process is examining the quality of the sequencing reads, performing quality trimming, and then checking the quality of the trimmed reads. It’s also…
Annotation - Olurida_v081 MAKER on Mox
2018
Olympia Oyster Genome Sequencing
Remarkably, I managed to burn through our Xsede computing resources and don’t have terribly much to show…
SRA Submission - Olymia oyster Whole Genome BS-seq Data
2018
Olympia oyster reciprocal transplant
Submitted our whole genome bisulfite sequencing data to NCBI Sequence Read Archive (SRA).
TrimGalore/FastQC/MultiQC - C.virginica Oil Spill MBDseq Concatenated Sequences
2018
LSU C.virginica Oil Spill MBD BS Sequencing
Previously concatenated and analyzed our Crassostrea virginica oil spill MBDseq data with FastQC./
Genome Annotation – Olympia oyster genome annotation results #02
2018
Olympia Oyster Genome Sequencing
Yesterday, I annotated our Olympia oyster genome using WQ-MAKER in just 7hrs!./
Genome Annotation - Olympia oyster genome annotation results #01
2018
Olympia Oyster Genome Sequencing
Yesterday, I annotated our Olympia oyster genome using WQ-MAKER in just 7hrs!./
Mox – Over quota: Olympia oyster genome annotation progress (using Maker 2.31.10)
2018
Olympia Oyster Genome Sequencing
Per this GitHub Issue, Steven has cleared out files.
Mox - Olympia oyster genome annotation progress (using Maker 2.31.10)
2018
Olympia Oyster Genome Sequencing
I decided to spend some time to figure…
Ubuntu - Fix “No Video Signal” Issue on Emu/Roadrunner
2018
Computer Servicing
Both Apple Xserves (Emu/Roadrunner)…
BS-seq Mapping - Olympia oyster bisulfite sequencing: TrimGalore > FastQC > Bismark
2018
BS-seq Libraries for Sequencing at Genewiz
MBD Enrichment for Sequencing at ZymoResearch
Steven asked me to evaluate our methylation sequencing data sets for Olympia oyster.
Assembly – SparseAssembler (k 111) on Geoduck Sequence Data
2018
Geoduck Genome Sequencing
Continuing to try to find the best kmer setting to work with SparseAssemlber after the last attempt failed due to a kmer size that was too large (k 131; which happens to be outside the max kmer size [127] for SparseAssembler), I re-ran SparseAssembler with an arbitrarily selected…
Data Management - Geoduck Phase Genomics Hi-C Data
2018
Geoduck Genome Sequencing
Samples Received
We received sequencing/assembly data from Phase Genomics.
Data Management - SRA Submission LSU C.virginica Oil Spill MBD BS-seq Data
2018
LSU C.virginica Oil Spill MBD BS Sequencing
Submitted the Crassostrea virginica (Eastern oyster) MBD BS-seq data we received on 20150413 to NCBI Sequence Read Archive./
TrimGalore/FastQC/MultiQC - Trim 10bp 5’/3’ ends C.virginica MBD BS-seq FASTQ data
2018
Miscellaneous
Steven found out that the Bismarck documentation (Bismarck is the bisulfite aligner we use in our BS-seq pipeline) suggests trimming 10bp from both the 5’ and 3’ ends. Since this is the next step in our pipeline, we figured we should probably just follow their…
TrimGalore/FastQC/MultiQC - 2bp 3’ end Read 1s Trim C.virginica MBD BS-seq FASTQ data
2018
Miscellaneous
Earlier today, I ran TrimGalore/FastQC/MultiQC on the Crassostrea virginica MBD BS-seq data from ZymoResearch and hard trimmed the first 14bp from each read. Things looked better at the 5’ end, but the 3’ end of each of…
Genome Assembly - SparseAssembler Geoduck Genomic Data, kmer=101
2018
Geoduck Genome Sequencing
Assembly complete. See end of post for data locations.