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This file is part of the "Universal Biomedical Skills" project.

Copyright (c) 2026 MD BABU MIA, PhD <md.babu.mia@mssm.edu>

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Finding Restriction Sites

Core Pattern

python
from Bio import SeqIO
from Bio.Restriction import EcoRI, BamHI, HindIII, RestrictionBatch, Analysis

record = SeqIO.read('sequence.fasta', 'fasta')
seq = record.seq

# Single enzyme
sites = EcoRI.search(seq)  # Returns list of cut positions

Search with Single Enzyme

python
from Bio.Restriction import EcoRI

sites = EcoRI.search(seq)
print(f'EcoRI cuts at positions: {sites}')
print(f'Number of sites: {len(sites)}')

# Check if enzyme cuts
if EcoRI.search(seq):
    print('EcoRI cuts this sequence')
else:
    print('EcoRI does not cut')

Search with Multiple Enzymes

python
from Bio.Restriction import RestrictionBatch, EcoRI, BamHI, HindIII, XhoI

batch = RestrictionBatch([EcoRI, BamHI, HindIII, XhoI])

# Method 1: batch.search()
results = batch.search(seq)
for enzyme, sites in results.items():
    if sites:
        print(f'{enzyme}: {sites}')

# Method 2: Analysis class
analysis = Analysis(batch, seq)
results = analysis.full()

Use Built-in Enzyme Collections

python
from Bio.Restriction import AllEnzymes, CommOnly

# All known enzymes (800+)
analysis = Analysis(AllEnzymes, seq)

# Commercially available only
analysis = Analysis(CommOnly, seq)

# Get results
results = analysis.full()
for enzyme, sites in results.items():
    if sites:
        print(f'{enzyme}: {sites}')

Linear vs Circular DNA

python
from Bio.Restriction import EcoRI, Analysis, RestrictionBatch

# Linear DNA (default)
sites_linear = EcoRI.search(seq, linear=True)

# Circular DNA (plasmid)
sites_circular = EcoRI.search(seq, linear=False)

# With Analysis class
batch = RestrictionBatch([EcoRI, BamHI])
analysis = Analysis(batch, seq, linear=False)  # Circular

Filter Results

python
from Bio.Restriction import Analysis, CommOnly

analysis = Analysis(CommOnly, seq)

# Only enzymes that cut
analysis.print_that_cut()

# Only enzymes that don't cut (non-cutters)
analysis.print_that_dont_cut()

# Enzymes that cut once
analysis.print_once_cutters()

# Enzymes that cut twice
analysis.print_twice_cutters()

# Get as dictionary
cutters = analysis.only_cut()
non_cutters = analysis.only_dont_cut()
once_cutters = analysis.once_cutters()
twice_cutters = analysis.twice_cutters()

Get Enzyme Information

python
from Bio.Restriction import EcoRI

# Recognition sequence
print(f'Site: {EcoRI.site}')           # GAATTC
print(f'Esite: {EcoRI.esite}')         # Recognition with cut position

# Cut characteristics
print(f'Overhang: {EcoRI.ovhg}')       # 4 (positive = 5' overhang)
print(f'Blunt: {EcoRI.is_blunt()}')    # False
print(f'5\' overhang: {EcoRI.is_5overhang()}')  # True
print(f'3\' overhang: {EcoRI.is_3overhang()}')  # False

# Overhang sequence
print(f'Overhang seq: {EcoRI.ovhgseq}')  # AATT

# Isoschizomers (same recognition, different cut)
print(f'Isoschizomers: {EcoRI.isoschizomers()}')

# Compatible enzymes (same overhang)
print(f'Compatible: {EcoRI.compatible_end()}')

Common Cloning Enzymes

python
from Bio.Restriction import (
    EcoRI, BamHI, HindIII, XhoI, SalI, NotI, XbaI, SpeI,
    NcoI, NdeI, BglII, PstI, KpnI, SacI, EcoRV, SmaI
)

common_enzymes = RestrictionBatch([
    EcoRI, BamHI, HindIII, XhoI, SalI, NotI, XbaI,
    NcoI, NdeI, BglII, PstI, KpnI, SacI, EcoRV, SmaI
])

analysis = Analysis(common_enzymes, seq)
results = analysis.full()

Access Enzymes by Name

python
from Bio.Restriction import AllEnzymes

# Get enzyme by string name
ecori = AllEnzymes.get('EcoRI')
sites = ecori.search(seq)

# Check if enzyme exists
if 'EcoRI' in AllEnzymes:
    print('EcoRI is in database')

Search Multiple Sequences

python
from Bio import SeqIO
from Bio.Restriction import RestrictionBatch, EcoRI, BamHI

batch = RestrictionBatch([EcoRI, BamHI])

for record in SeqIO.parse('sequences.fasta', 'fasta'):
    analysis = Analysis(batch, record.seq)
    results = analysis.full()
    print(f'{record.id}:')
    for enzyme, sites in results.items():
        if sites:
            print(f'  {enzyme}: {sites}')

Notes

• Positions are 1-based - first base is position 1
• Cut position - where enzyme cuts (between bases)
• Linear default - set linear=False for circular DNA
• Case insensitive - recognition matches regardless of case
• Ambiguous bases - some enzymes recognize N, R, Y, etc.

Related Skills

• restriction-mapping - Visualize cut positions on sequence
• enzyme-selection - Choose enzymes by criteria
• fragment-analysis - Analyze resulting fragments

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