Sequence I/O

When to Use This Skill

• Reading or writing sequence files (FASTA, GenBank, FASTQ)
• Converting between sequence file formats
• Manipulating sequences (complement, reverse complement, translate)
• Extracting sequences from large indexed FASTA files (faidx)
• Calculating sequence statistics (GC content, molecular weight, Tm)

When NOT to Use This Skill

• NGS alignment files (SAM/BAM/VCF) → Use

  pysam

• BLAST searches → Use

  gget

  (quick) or

  blat-integration

  (large-scale)
• Multiple sequence alignment → Use

  msa-advanced

• Phylogenetic analysis → Use

  etetoolkit

• NCBI database queries → Use

  pubmed-database

  or

  gene-database

Tool Selection Guide

Bio.SeqIO

biopython_seqio.md

Bio.SeqIO.convert()

biopython_seqio.md

Bio.Seq

biopython_seqio.md

pysam.FastaFile

faidx.md

Bio.SeqUtils

utilities.md

Quick Start

Installation

uv pip install biopython pysam

Read FASTA

from Bio import SeqIO

for record in SeqIO.parse("sequences.fasta", "fasta"):
    print(f"{record.id}: {len(record.seq)} bp")

Convert GenBank to FASTA

from Bio import SeqIO

SeqIO.convert("input.gb", "genbank", "output.fasta", "fasta")

Random Access with faidx

import pysam

# Create index (once)
pysam.faidx("reference.fasta")

# Random access
fasta = pysam.FastaFile("reference.fasta")
seq = fasta.fetch("chr1", 1000, 2000)  # 0-based coordinates
fasta.close()

Sequence Operations

from Bio.Seq import Seq

seq = Seq("ATGCGATCGATCG")
print(seq.complement())
print(seq.reverse_complement())
print(seq.translate())

Reference Documentation

references/biopython_seqio.md

• Bio.Seq

  object and sequence operations

• Bio.SeqIO

  for file parsing and writing

• SeqRecord

  object and annotations
• Supported file formats
• Format conversion patterns

references/faidx.md

• Creating FASTA index with

  pysam.faidx()

• pysam.FastaFile

  for random access
• Coordinate systems (0-based vs 1-based)
• Performance considerations for large files
• Common patterns (variant context, gene extraction)

references/utilities.md

• GC content calculation (

  gc_fraction

  )
• Molecular weight (

  molecular_weight

  )
• Melting temperature (

  MeltingTemp

  )
• Codon usage analysis
• Restriction enzyme sites

references/formats.md

• FASTA format specification
• GenBank format specification
• FASTQ format and quality scores
• Format detection and validation

Coordinate Systems

• Numeric arguments: 0-based (

  fetch("chr1", 999, 2000)

  = positions 999-1999)
• Region strings: 1-based (

  fetch("chr1:1000-2000")

  = positions 1000-2000)

Common Pitfalls

1. Coordinate confusion: Remember which tool uses 0-based vs 1-based
2. Missing faidx index: Random access requires

   .fai

   file
3. Format mismatch: Verify file format matches the format string in

   SeqIO.parse()

4. Iterator exhaustion:

   SeqIO.parse()

   returns an iterator; convert to list if multiple passes needed
5. Large files: Use iterators, not

   list()

   , for memory efficiency