GSE181137 Processing Pipeline

Publication

Cell reports (2022) — PMID 35263585

Dataset

Identification of the Global miR-130a Targetome Reveals a Novel Role for TBL1XR1 in Hematopoietic Stem Cell Self-Renewal and t(8;21) AML [miR130aKD_K…

1. 1

   Raw Reads were aligned using STAR against known GENCODE 32 transcripts

   STAR v2.7.10a (Inferred with models/gemini-2.5-flash)

   $ Bash example

   # --- Installation (example using Conda) ---
   # conda create -n star_env star=2.7.10a -y
   # conda activate star_env
   
   # --- Reference Data Setup (GENCODE 32 for GRCh38) ---
   # This section demonstrates how to prepare the STAR genome index for GENCODE 32.
   # The actual alignment command assumes this index is already built.
   
   # # Create a directory for the genome index
   # mkdir -p star_gencode32_index
   # cd star_gencode32_index
   
   # # Download GENCODE 32 human genome (GRCh38 primary assembly) FASTA and GTF files
   # wget -c ftp://ftp.ebi.ac.uk/pub/databases/gencode/Gencode_human/release_32/GRCh38.primary_assembly.genome.fa.gz
   # wget -c ftp://ftp.ebi.ac.uk/pub/databases/gencode/Gencode_human/release_32/gencode.v32.annotation.gtf.gz
   
   # # Unzip the downloaded files
   # gunzip GRCh38.primary_assembly.genome.fa.gz
   # gunzip gencode.v32.annotation.gtf.gz
   
   # # Build the STAR genome index
   # # Adjust --sjdbOverhang based on the maximum read length minus 1 (e.g., 74 for 75bp reads).
   # # For eCLIP, read lengths can vary, often 50-75bp. Using 74 as a common example.
   # STAR --runMode genomeGenerate \
   #      --genomeDir . \
   #      --genomeFastaFiles GRCh38.primary_assembly.genome.fa \
   #      --sjdbGTFfile gencode.v32.annotation.gtf \
   #      --sjdbOverhang 74 \
   #      --runThreadN 8 # Adjust number of threads as needed
   # cd ..
   
   # --- Alignment Step ---
   # Define variables for input and output
   GENOME_DIR="star_gencode32_index" # Path to your pre-built STAR genome index
   READS_R1="raw_reads_R1.fastq.gz" # Placeholder for input FASTQ file (Read 1)
   READS_R2="raw_reads_R2.fastq.gz" # Placeholder for input FASTQ file (Read 2, remove for single-end)
   OUTPUT_PREFIX="aligned_sample" # Prefix for output files (e.g., aligned_sample.Log.out, aligned_sample.Aligned.sortedByCoordinate.out.bam)
   NUM_THREADS=8 # Adjust number of threads as needed
   
   # Execute STAR alignment
   STAR --genomeDir "${GENOME_DIR}" \
        --readFilesIn "${READS_R1}" "${READS_R2}" \
        --readFilesCommand zcat \
        --runThreadN "${NUM_THREADS}" \
        --outFileNamePrefix "${OUTPUT_PREFIX}." \
        --outSAMtype BAM SortedByCoordinate \
        --outSAMattributes All \
        --outFilterMultimapNmax 20 \
        --outFilterMismatchNmax 999 \
        --outFilterMismatchNoverLmax 0.04 \
        --alignIntronMin 20 \
        --alignIntronMax 1000000 \
        --alignMatesGapMax 1000000 \
        --sjdbScore 1 \
        --quantMode GeneCounts # Optional: Output gene counts (ReadsPerGene.out.tab)

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Tools Used