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FreeGenes Project Open Enzyme Collection


DNA polymerase structure
Open Bioeconomy Labs logo

The Open Enzyme Collection consists of more than 75 genes for enzymes that are considered "workhorses" of molecular biology, including DNA polymerases, RNA polymerases, DNA ligases, reverse transcriptases, and restriction enzymes. These enzymes are useful in both basic and applied fields as diverse as genomics, diagnostics, biodiversity, synthetic biology, DNA origami, immunology, biochemistry, and more. The foundational technologies enabled by this collection include polymerase chain reaction (PCR), cloning, isothermal amplification, reverse transcription of RNA into DNA, and dNTP synthesis. Additionally, the Open Enzyme collection contains enzyme genes for fixing damaged and degraded DNA prior to amplification, enzymes for cutting protein sequences to remove unwanted parts, and DNA-binding proteins that can be fused with other enzymes to improve functionality. The Open Bioeconomy Lab designed and codon-optimized the Open Enzymes parts for expression in Escherichia coli bacteria. Researchers can use the collection as the basis of manufacturing their own enzymes by cloning them into an expression vector of their choice and transforming into E. coli bacteria. This can help reduce lab costs and overcome supply chain issues, particularly in areas of the world where enzymes are harder to obtain.

The enzymes can also be used as a base to improve functionality or add new functions through genetic engineering and directed evolution. More information about the collection can be obtained at the Open Bioeconomy Lab's Open Enzyme Collection.

How to Use

The plasmids in the Open Enzyme collection contain genetic parts and are not functional by themselves. The selection marker for all plasmids in this collection is ampicillin. Find more information on the enzymes and protocols for expression at Open Bioeconomy Lab's crowd-sourced expression guides. Researchers can clone these enzymes into an expression vector of their choice and then transform into E. coli bacteria for expression and purification (pET28a-LIC or other pET vectors are recommended). Open Enzymes are also compatible with the E. coli Protein Expression Toolkit (all Open Enzyme parts have C/E overhangs for Golden Gate assembly).


Explore the collection and order individual plasmids via the links below:

DNA Ligases

Addgene ID Plasmid Gene/insert
165503 pOpen-HSlig4 DNA Ligase IV
165514 pOpen-9N7lig 9 Degrees North DNA Ligase
165525 pOpen-Pfulig Pfu DNA Ligase
165535 pOpen-THEAQlig Taq DNA Ligase
165554 pOpen-T4gene30 T4-DNA Ligase
165564 pOpen-ECOligA E. coli DNA Ligase
165569 pOpen-Tthlig Tth DNA Ligase
165574 pOpen-HSlig3 DNA Ligase III

DNA Polymerases

Addgene ID Plasmid Gene/insert
165500 pOpen-9N7polA 9 degrees N-7 DNA polymerase
165501 pOpen-GBDpol Pyrococcus sp. Heat-Stable DNA Polymerase
165502 pOpen-9N7polA (CT) 9 degrees N-7 DNA Polymerase chain terminating
165511 pOpen-K12polLF DNA Polymerase I, Large (Klenow) Fragment
165522 pOpen-K12polLF (exo-) DNA Polymerase I, Large (Klenow) Fragment (3'-5' exo-)
165512 pOpen-THEAQpolA Taq DNA Polymerase
165513 pOpen-dbh Sulfolobus DNA Polymerase IV
165523 pOpen-Tlipol (exo-) Tli DNA polymerase (exo-)
165524 pOpen-T4gene43 T4 DNA Polymerase
165532 pOpen-ECOpolA E. coli DNA Polymerase I
165533 pOpen-TthpolA Tth DNA polymerase
165534 pOpen-T7gene5 T7 DNA Polymerase (unmodified)
165542 pOpen-KTHEAQpol KlenTaq1 polymerase
165543 pOpen-Bstpol Bst DNA Polymerase, Full Length
165544 pOpen-T5gene122 T5 DNA Polymerase
165553 pOpen-BstpolLF Bst DNA Polymerase, Large Fragment
165562 pOpen-Pwopol Pwo DNA Polymerase
165563 pOpen-BsupolLF Bsu DNA Polymerase I, Large Fragment
165570 pOpen-P45 P45
165572 pOpen-GBDpol (exo) Pyrococcus sp. Heat-Stable (exo-) DNA Polymerase
165573 pOpen-phi29pol phi29 DNA Polymerase

Restriction Endonucleases

Addgene ID Plasmid Gene/insert
165504 pOpen-EcoRIR EcoRI
165506 pOpen-Eco31I Eco31I
165507 pOpen-NotIR (GC) NotI (GC-rich recognition site)
165508 pOpen-HindIIIR HindIII
165526 pOpen-PstIR PstI
165528 pOpen-SfiIR (cDNA) SfiI (used to create cDNA libraries)
165529 pOpen-DpnIR DpnI
165537 pOpen-SapIR SapI
165539 pOpen-Bst HF Bst HF
165548 pOpen-EcoRV EcoRV
165567 pOpen-XbaIR XbaI
165568 pOpen-NcoIR NcoI

Reverse Transcriptases

Addgene ID Plasmid Gene/insert
165546 pOpen-MMLV_RT (mut H) Moloney Murine Leukemia Virus (MMLV) Reverse Transcriptase (RNAse H deactivated by 3 mutations)
165556 pOpen-MMLV_RT (lack H) Moloney Murine Leukemia Virus (MMLV) Reverse Transcriptase RNaseH - (lacking RNaseH domain)
165566 pOpen-TthRT Thermus thermophilus (Tth) Reverse Transcriptase

RNA Polymerases

Addgene ID Plasmid Gene/insert
165505 pOpen-T7gene1 T7 RNA Polymerase
165516 pOpen-SP6rpol SP6 RNA Polymerase
165576 pOpen-T3gene1 T3 RNA Polymerase

Other Enzymes

Addgene ID Plasmid Gene/insert
165509 pOpen-TEV protease TEV protease
165565 pOpen-RNAse A RNAse A
165575 pOpen-RNAse H RNAse H
165510 pOpen-DNA Topoisomerase 1B Vaccinia Virus Vaccinia Virus DNA Topoisomerase 1B
165515 pOpen-EcoRIM M.EcoRI DNA methyltransferase
165517 pOpen-Eco31IA Eco31IA DNA methyltransferase
165518 pOpen-NotIM M.NotI DNA methyltransferase
165527 pOpen-Eco31IB Eco31IB DNA methyltransferase
165536 pOpen-PstIM M.PstI DNA methyltransferase
165538 pOpen-SfiIM M.SfiI 165538 DNA methyltransferase
165547 pOpen-SapIM1 M1.SapI DNA methyltransferase
165557 pOpen-SapIM2 M2.SapI DNA methyltransferase
165558 pOpen-EcoRVM M.EcoRV DNA methyltransferase
165577 pOpen-XbaIM M.XbaI DNA methyltransferase
165578 pOpen-NcoIM M.NcoI DNA methyltransferase
165520 pOpen-T4 PNK T4 PNK polynucleotide kinase
165521 pOpen-lambda red operon lambda red operon
165530 pOpen-T4-BGT T4-BGT beta-glucosyltransferase
165531 pOpen-UDG UDG Uracil DNA glycosylase
165541 pOpen-Fpg Fpg Formamidopyrimidine DNA glycosylase
165550 pOpen-T4 PGD T4 PGD (T4 pyrimidine DNA glycosylase, aka T4 Endonuclease V)
165540 pOpen-T7EI T7 Endonuclease I
165551 pOpen-Endonuclease IV E. coli E. coli Endonuclease IV
165561 pOpen-Endonuclease VIII E. coli E. coli Endonuclease VIII
165571 pOpen-Exonuclease III E. coli E. coli Exonuclease III
165559 pOpen-T5gene12/13 T5 Exonuclease
165560 pOpen-T4 g32 T4 gene 32 single-stranded DNA-binding protein
165549 pOpen-ETSSB Extreme Thermostable Single-Stranded DNA-Binding Protein
165579 pOpen-dromedNK dNK from Drosophila
165580 pOpen-BovDNTT Bovine DNTT
165555 pOpen-ALPI CIP (calf intestinal phosphatase)

Additional Resources