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| | miniGENES (up to 400 bp) | Genes (401 + bp) |
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| Pricing | $220.00 USD | Starting at $0.55 USD / bp | | Turnaround Time | 4-8 business days | 8-12 business days |
| | *The time it takes to make a gene is dependent on length, complexity, and vector choice. | | *Extra charges may apply for gene segments with added complexity (such as homopolymeric runs, critical hairpin structures or G/C content), which can interfere with assembly and/or sequencing performance. For more information about potential types of complexity, please see the Overview section below. | |
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- Design review and consultation
- Researchers enter the gene sequence on the gene entry site.
- A/C/G/T bases only
- no mixed degenerate bases
- no “modified" bases (Inosine, Uridine)
- The sequence is automatically analyzed for the following characteristics which may interfere with synthesis, assembly, or sequencing performance:
- Hairpins: Hairpins >10 bases and G/C-rich secondary structures
- Repeats and homo-polymeric runs: long repeats, >10 bases for G/C, or >30 bases for A/T
- Overall and regional GC content: content >65% and <25% for G/C
- Restriction Site Analysis: restriction site duplications and Dam/Dcm Methylation sites
- Dam Methylation Site = GmATC
- Dcm Methylation Site = CmCWGG
- If the sequence does not pass the screen criteria, you will be contacted by a gene services specialist to determine the best way to proceed.
- The sequence will also be analyzed to ensure safety and regulatory conformance.
- A biohazard disclosure statement is required for each gene order.
- IDT reserves the right to refuse any order that does not pass this analysis.
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| Different organisms vary in the frequency with which they use certain codons and exhibit varying degrees of codon preferences. The fastest growing microbes tend to have the highest codon biases. This codon bias is thought to be reflective of the abundance of the corresponding tRNA pool. Gene sequences containing preferred codons for the expression host are translated more efficiently, with greater accuracy, and in greater yield. In addition, optimization can be used to disrupt difficult sequence features (such GC content, hairpins, repeats, etc.). A codon optimization tool is available on the IDT website. | |
| Once a gene is cloned, IDT verifies the sequence with double-stranded DNA sequencing. We also provide the sequencing chromatograms and a plasmid map to the customer. | |
| IDT will provide 2 µg of purified double-stranded DNA in a circularized plasmid, delivered lyophilized. | |
- All sequence information is confidential at IDT
- All online ordering steps, including sequence entry and choice of parameters, is secure and protected.
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| IDT provides the gene in a cloning vector so that it is amendable to further subcloning into a vector of the researcher’s choice. These genes can then be used in a number of applications including, but not limited to: | - Protein Expression
- Recombinant antibodies
- Novel fusion proteins
- Codon optimized short proteins
- Functional peptides: catalytic, regulatory, binding domains
- microRNA genes
- Template for in vitro transcription (IVT)
- shRNA expression cassettes
- Regulatory sequence cassettes
- Synthetic repeat constructs
- Synthetic chromosomes/genomes
- Custom vectors
- Microarray-ready cDNA
- Gene variants and SNPs
- DNA vaccines and vectors
- Standards for quantitative PCR and other assays
- Functional Genomics
- Custom genes offer near limitless flexibility for protein mutagenesis
- Unrestricted point mutations
- Mutant libraries
- Large deletion mutants
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| IDT has the ability to clone in-house in a range from 50bases -5.0kb. All genes, unless otherwise specified, will be delivered in IDT’s proprietary cloning vector, pIDTSmart. This vector has been specifically engineered to remove most common restriction endonuclease cleaving sites and does not contain a promoter within the cloning region. If you require restriction sites or promoters for your application, it is important to include these when ordering the gene of interest by appending the desired restriction sites or promoters to the ends of your gene construct. The default version of pIDTSmart contains a kanamycin resistance cassette. An ampicillin version is also available upon request. | |
| IDT has the ability to sub-clone into almost any customer supplied vector. To get the process started, go to IDT’s website (www.idtdna.com/order/customvector.aspx) and complete the Custom Vector request sheet or contact IDT Gene Support at genes@idtdna.com. An IDT representative will contact you within 2 business days to discuss your custom vector request. |
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| pIDTBlue | ApaLI | GTGCAC | 2 | | | BspDI | ATCGAT | 1 | | | ClaI | ATCGAT | 1 | | | EagI | CGGCCG | 1 | Doesn't Exist in <= 400 version | | PvuI | CGATCG | 3 | | | PvuII | CAGCTG | 2 | | | RsaI | GTAC | 2 | | | Restriction Sites not Present in Plasmid | | Acc65I, AgeI, ApaI, AscI, Asp718I, AvaI, AvrII, BamHI, BglII, BmgBI, BstBI, DraII, Ecl136II, EcoRI, EcoRV, HindIII, HpaI, KpnI, MluI, NcoI, NdeI, NheI, NotI, NsiI, PacI, PaeR7I, PspOMI, PstI, SacI, SacII, SalI, SmaI, SnaBI, SpeI, SphI, TliI, TspMI, XbaI, XhoI, XmaI |
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| pIDTSmart-KAN/AMP | ApaL | GGGCCC | 1 | Doesn't Exist in > 400 version or pIDTSMART-KAN | | ApaLI | GTGCAC | 2 | Contains 1 in pIDTSMART-KAN | | BspDI | ATCGAT | 1 | Contains 2 in pIDTSMART-KAN | | ClaI | ATCGAT | 1 | Contains 2 in pIDTSMART-KAN | | DraII | RGGNCCY | 2 | Contains 1 in pIDTSMART-KAN | | NsiI | ATGCAT | 2 | Doesn't Exist in <= 400 version | | PspOMI | GGGCCC | 1 | Doesn't Exist in > 400 version or pIDTSMART-KAN | | PvuI | CGATCG | 3 | | | RsaI | GTAC | 2 | | | SnaBI | TACGTA | 1 | | | Restriction Sites not Present in Plasmid | | Acc65I, AgeI, AscI, Asp718I, AvaI, AvrII, BamHI, BglII, BmgBI, BstBI, EagI, Ecl136II, EcoRI, EcoRV, HindIII, HpaI, KpnI, MluI, NcoI, NdeI, NheI, NotI, PacI, PaeR7I, PstI, PvuII, SacI, SacII, SalI, SmaI, SpeI, SphI, TliI, TspMI, XbaI, XhoI, XmaI |
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- Centrifuge the tube prior to opening to ensure that the DNA is at the bottom of the tube.
- Resuspend DNA in 20 µL of 10 mM Tris, 0.1 mM EDTA buffer (pH 7.5 – 8.0) to reach an approximate concentration of 0.1 µg/µL for a stock concentration.
- Incubate the tube at room temperature for 30 minutes and then vortex for 20 seconds
- Centrifuge for 1 minute
- To create a working concentration, add 1 µL of the stock concentration to 999 µL of water to reach an approximate working concentration of 0.1 ng/µL. Use 1-2 µL of the working dilution for bacterial transformations.
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Store hydrated DNA at -20°C.
Lyophilized DNA is stable at room temperature for 3 months. We recommend storing at -20°C for longer periods of time. | | Transformations (Chemical) | - Place frozen competent cells and pre-labeled tubes on ice
- Pre-warm the hot bath to 42°C
- Once cells are thawed on ice, aliquot 25 µL of XL1 Blue cells to each of the pre-chilled tubes. Keep all the tubes on ice
- Add 2 µL of ligation or PCR reaction to cells and gently swirl the cells with pipette tip
- Incubate on ice for 30 minutes
- Heat shock tubes for 45 seconds in 42°C hot bath
- Return tubes to ice for 2 minutes
- Add 250 µL of SOC Media to each tube and place tubes in a 37°C shaking incubator for 1 hour
- Pre-warm agar plates to 37°C in incubator
- Spread 125 µL of cells on agar plates
- After spreading cells, let plates sit for approximately 10 minutes at room temperature. Then place the plates upside down in 37° incubator for 12 – 24 hours
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- For a large number of colonies, use a 2.2 mL deep well plate and add 1.6 mL of LB Broth with the appropriate antibiotic.
For a small number of colonies, use a 14 mL round bottom culture tube and add 2 mL of LB Broth with the appropriate antibiotic - Touch the colony with a toothpick or pipette tip and place it in the broth
- Cover with a gas permeable lid and place in a 37° shaking incubator for 12 – 20 hours.
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| The standard in-house digestion protocol is below: | | DNA | 400ng | | Buffer | 5µL | | Water | XµL | | Enzyme(s) | 1µL each | | Total Volume | 50µL |
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| | Incubate for 1 hour at the temperature recommended by the enzyme manufacturer. Take 2-5 µL of the digested sample, add loading dye, and run on a gel to verify that the digestion took place. In separate lanes, also run a ladder to verify the size of the product and a sample of undigested product as a control. | | *The length of running time for the gel will depend on the size of the digested insert. Smaller inserts require shorter run times or the product will run off the gel. | | *The enzyme should not be more than 1/10th of total reaction | |
| Material | Components | Amount (g or ml)/L |
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| STE Buffer pH 7.5 | 10 mM | Tris | 1.21 | | 10 mM | NaCl | 2.92 | | 1 mM | EDTA | 0.37 | | 10X Taq Ligase Buffer pH 7.6 | 200 mM | Tris | 24.22 | | 250 mM | Potassium Acetate | 24.54 | | 100 mM | Magnesium Acetate (tetrahydrate) | 21.45 | | 100 mM | DTT | 15.42 | | 100 mM | NAD | 6.85 | | 1 % | Triton-X100 | 24.22 | | 1X T4 DNA Ligase Buffer pH 7.5 | 50 mM | Tris | 6.05 | | 10 mM | Magnesium Acetate (tetrahydrate) | 2.03 | | 10 mM | DTT | 1.54 | | 1 mM | Adenosine 5′-triphosphate disodium salt | 0.55 | | 25 µg/ml | BSA | 0.025 | | Agar Plates with Amp | 1 % | Tryptone | 10 | | 1 % | Sodium Chloride | 10 | | 0.5 % | Yeast Extract | 5 | | 1.5 % | Agar | 15 | | 100 µg/ml | Ampicillin | 0.1 | | LB Broth with Amp | 1 % | Tryptone | 10 | | 1 % | Sodium Chloride | 10 | | 0.5 % | Yeast Extract | 5 | | 100 µg/ml | Ampicillin | 0.1 | | Agar Plates with Kan | 1 % | Tryptone | 10 | | 1 % | Sodium Chloride | 10 | | 0.5 % | Yeast Extract | 5 | | 1.5 % | Agar | 15 | | 50 µg/ml | Kanamycin | 0.05 | | LB Broth with Kan | 1 % | Tryptone | 10 | | 1 % | Sodium Chloride | 10 | | 0.5 % | Yeast Extract | 5 | | 50 µg/ml | Kanamycin | 0.05 | | SOC Media pH 7.5 | 2 % | Tryptone | 20 | | 0.5 % | Yeast Extract | 5 | | 0.05 % | Sodium Chloride | 0.5 | | 2.5 mM | Potassium Chloride | 0.19 | | 10 mM | Magnesium Chloride (hexahydrate) | 20.3 | | 1X Loading Dye | 0.4 % | Bromophenol Blue | 4 | | 0.4 % | Xylene Cyanol FF | 4 | | 50 % | Glycerol | 500 |
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As one of five founding members of the International Gene Synthesis Consortium (IGSC), IDT’s gene sequence and customer screening practices are consistent with the IGSC’s Harmonized Screening Protocol. To prevent the misuse of synthetic genes, IDT screens the sequences of ordered genes to identify regulated and other potentially dangerous pathogen sequences, and verifies that its customers are legitimate scientists engaged in beneficial research.
For more information about the IGSC and the Harmonized Screening Protocol, please visit the website at http://www.genesynthesisconsortium.org/Home.html.
The United States government has published draft guidance describing how commercial providers of synthetic genes should perform gene sequence and customer screening. IDT, along with the other IGSC member companies, supports the adoption of that guidance in substance and will implement the procedures that the final guidance recommends. For more information, please see http://edocket.access.gpo.gov/2009/E9-28328.htm.
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