FreeGenes Project Open Enzyme Collection
- DNA Ligases
- DNA Polymerases
- Restriction Endonucleases
- Reverse Transcriptases
- RNA Polymerases
- Other Enzymes
Background
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).
Plasmids
Explore the collection and order individual plasmids via the links below:
- DNA Ligases
- DNA Polymerases
- Restriction Endonucleases
- Reverse Transcriptases
- RNA Polymerases
- Other Enzymes
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) |