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Store at -20°C. For in-vitro laboratory use only

General Description

Bullseye Taq DNA Polymerase is a thermostable recombinant DNA polymerase, which exhibits very high activity in primer extension and other molecular biology applications. The enzyme is isolated from Thermus aquaticus and has a molecular weight of approximately 94 kDa.

Bullseye Taq DNA Polymerase has both a 5'®3' DNA polymerase and a 5'®3' exonuclease activity. The enzyme lacks a 3'®5' exonuclease activity (no proofreading ability). Taq DNA Polymerase leaves an A¢ overhang, which makes the enzyme ideal for TA cloning.

The 10X Reaction Buffer provided does not contain Mg+2. 25 mM MgCl2 is supplied separately.

10X Mg++ Free Standard Buffer

100 mM Tris-HCl pH 8.3, 500 mM KCl.

Unit Definition

One unit is defined as the amount that incorporates 10 nmoles of dNTPs into acid-precipitable form in 30 minutes at 72°C under standard assay conditions.

Storage and Dilution Buffer

Enzyme is supplied in 20 mM Tris-HCl pH 8.3, 100 mM KCl, 0.1 mM EDTA, 1 mM DTT, 0.5% Tween 20, 0.5%NP40, 50% glycerol.

Quality Control

Each lot of Taq DNA Polymerase is tested for contaminating activities, with no trace of endonuclease activity, nicking activity, exonuclease activity or priming activity.

Suggested Protocol using Taq DNA Polymerase

This protocol serves as a guideline only. Optimal reaction conditions may vary and must be individually determined.

Table 1. Reaction components (master mix & template DNA)

Table 2. MgCl2 concentration in a 50 mL reaction

Table 2 provides the volume of 25 mM MgCl2 to add to the master mix if a certain MgCl2 concentration is required.

2x Master Mix Kit (1.5 mM MgCl2) 

Bullseye Taq DNA Polymerase Mix is a ready-to-use 2x reaction mix. Simply add primers, template, and water to successfully carry out primer extensions and other molecular biology applications.

Bullseye Taq polymerase, the NH4+ buffer system, dNTPs, and magnesium chloride are conveniently present in the Taq DNA Polymerase Mix. (Inert Red Dye is present in BE180303 only)

Bullseye Taq DNA Polymerase Mix offers several advantages. Set up time is significantly reduced. The chance of contaminating component stocks is eliminated. Reduction of reagent handling steps leads to better reproducibility. Standard tests can be set up with the confidence that results will be consistent every time.

Composition of 2x Taq Master Mix

150 mM Tris-HCl pH 8.5, 40 mM (NH4)2S04, 3 mM MgCl2, 0.2% Tween 20Ò

4 mM dNTPs

2 units/µL AS ONE Taq polymerase

Inert Red Dye & Stabilizer (BE180303 only)

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New and Improved! Validated for SNP Genotyping assays, Gene expression assays, Microarray validation and High throughput screening!

Bullseye EvaGreen qPCR MasterMix is designed for quantitative/real-time analysis. The components of Bullseye EvaGreen qPCR MasterMix has been developed for superb performance in sensitivity, signal-to-noise ratio, and complete elimination of primer dimers. The chemically modified Hot-Start Taq polymerase, included in our Mastermix, significantly reduces non-specific PCR amplification observed with regular Taq polymerase. Every lot is tested on different real-time PCR machines listed below.

• Instrument-specific & pre-optimized
• Validated applications
• Reproducible results
• Highly competitive pricing

For laboratory research only. Not for clinical applications.

For technical questions, phone the MIDSCI helpline at 800-227-9997

• Inert yellow dye helps reduce pipetting errors
• Room temperature stable for up to 30 days
• Compatible with fast cycling protocols
• Highly sensitive for low copy number templates
• Includes PR1MA Hot Start Taq Polymerase

Successful PCR requires careful control of many variables.  Pipetting errors, poor performing polymerases, dNTP concentrations, are just a few of the variables that can all contribute to reaction problems.  PR1MA has developed their new qMAX™ Gold to control these variables and help you achieve the best possible amplification performance. To reduce the chance of pipetting errors, qMAX™ Gold includes an inert yellow dye so small volumes are easy to visualize in PCR plates.  

qMAX™ Gold is a ready to use, 2X mastermix of PR1MA Hot Start Taq enzyme, dNTPs, a sensitive fluorescent intercalating dye, and optimized reaction buffer.  The Hot Start Taq allows reaction set up at room temperature while the temperature stable formulation guarantees optimal performance even if the mix is left at room temperature for extended periods.

Just add primers and DNA targets to the mix, and then proceed to amplification.  qMAX™ Gold is compatible with all real time thermal cyclers and exhibits high sensitivity with normal or fast 2-step cycling protocols.
 *Please note these products ship on dry ice.  Appropriate shipping charges apply unless otherwise noted on a quote.
 
 

Cas9 Nuclease 
Clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated (Cas) systems provide bacteria and archaea with adaptive immunity against viruses and plasmids by using CRISPR RNAs (crRNAs) to guide the silencing of invading nucleic acids. The CRISPR system consists of a short non-coding guide RNA (sgRNA) made up of a target complementary CRISPR RNA (crRNA) and an auxiliary transactivating crRNA (tracrRNA). The sgRNA guides the Cas9 endonuclease to a specific genomic locus via base pairing between the crRNA sequence and the target sequence, and cleaves the DNA to create a double-strand break. The location of the break is within the target sequence 3 bases from the NGG PAM (Protospacer Adjacent Motif). The PAM sequence, NGG, must follow the targeted region on the opposite strand of the DNA with respect to the region complementary sgRNA sequence (Fig.1).

Intact Genomics Cas9 Nuclease is the purified recombinant Streptococcus pyogenes Cas9 enzyme containing a nuclear localization signal (NLS) at the C-terminal for targeting to the nucleus. This enzyme is designed to perform CRISPR/Cas9-mediated genome editing. The physical purity of this enzyme is â¥98% as assessed by SDS-PAGE with Coomassie® blue staining.

Quality Control 
Quality control is performed following the production of each new lot of product to ensure that it meets the quality standards and specifications designated for the product. Each lot is repeatedly compared side-by-side with leading competitors to ensure our products outperform the competitor before product launching. 

Cas9 nuclease is free from detectable RNase, Endonuclease (nicking) and non-specific DNase activities.

Product Source 
E. coli BL21 (DE3) strain expressing a Cas9 gene from Streptococcus pyogenes with an N-terminal 6xHis tag and C-terminal SV40 nuclear localization signal (NLS).

Contents & Storage

1. Cas9 Nuclease
2. 10x Cas9 Nuclease Reaction Buffer

Store Cas9 Nuclease and Buffer at -20 °C

Storage Buffer 
50 mM Tris-HCl, 50 mM KCl, 1 mM DTT, 0.1 mM EDTA, 50% Glycerol, pH 7.5 @ 25 °C

1x Cas9 Reaction Buffer 
20 mM HEPES, 100 mM NaCl, 5 mM MgCl, 0.1 mM EDTA, pH 6.5 @ 25 °C

Functional Testing
Cas9 Nuclease functional testing was done by in vitro DNA cleavage assay with the following protocol which gives more than 95% digestion of the substrate DNA as determined by agarose gel electrophoresis.
1) Set up 30 µl reaction in a microcentrifuge tube on ice with the following combinations.

T4 DNA Ligase

Intact Genomics T4 DNA Ligase catalyzes the formation of a phosphodiester bond between juxtaposed 5'-phosphate and 3'-hydroxyl termini in duplex DNA or RNA.  This enzyme joins DNA fragments with either cohesive or blunt termini as well as repair single stranded nicks in duplex DNA, RNA or DNA/RNA hybrids.

Intact Genomics T4 DNA Ligase catalyzes the formation of a phosphodiester bond between juxtaposed 5'-phosphate and 3'-hydroxyl termini in duplex DNA or RNA.  This enzyme joins DNA fragments with either cohesive or blunt termini as well as repair single stranded nicks in duplex DNA, RNA or DNA/RNA hybrids.

Quality Control 
Quality control is performed following the production of each new lot of product to ensure that it meets the quality standards and specifications designated for the product. Each lot is repeatedly compared side-by-side with leading competitors to ensure our products outperform the competitor before product launching. 

Intact Genomics T4 DNA Ligase displays up to 3-5X higher ligation efficiency than the nearest competitor.

Product Source
E. coli strain expressing a recombinant clone

Quality Control 
Quality control is performed following the production of each new lot of product to ensure that it meets the quality standards and specifications designated for the product. Each lot is repeatedly compared side-by-side with leading competitors to ensure our products outperform the competitor before product launching. 

Contents & Storage

1. T4 DNA Ligase
2. 10x T4 DNA Ligase Reaction Buffer (w/o ATP)
3. 10 mM ATP

Store all contents at -20 °C.

Storage Buffer 
50 mM Tris-HCl, 50 mM KCl, 1 mM DTT, 0.1 mM EDTA, 50% Glycerol, pH 7.5 @ 25 °C

10x T4 DNA Ligase Reaction Buffer (w/o ATP) 
500 mM Tris-HCl, 100 mM MgCl, 100  mM DTT, pH 7.5 @ 25 °C

Note
10x T4 DNA ligase buffer does not contain ATP. You need to add ATP separately.

Unit Definition 
One Weiss unit is defined as the amount of enzyme required to convert 1 nmol of 32P from pyrophosphate into Norit-absorbance material in 20 minutes under standard assay conditions.

Protocol

1. Set up reaction buffer in a microcentrifuge tube on ice. Use a molar ratio of 1:3 vector to insert DNA.

1. Gently mix the reaction and centrifuge briefly.
2. For cohesive ends, incubate 16 °C for overnight or at room temperature for 30 min.
3. For blunt ends, incubate 16 °C for overnight or at room temperature for 2 hrs.
4. Heat inactivate at 70 °C for 15 min.
5. Cool on ice and transform 2 µl of the reaction into 50 µl competent cells.

ig-Fusion Cloning Kit 
Intact Genomics propriety ig-Fusion cloning technology is a simple, rapid and highly efficient cloning kit which allows to directly clone any PCR product(s) to any linearized expression vector at any site. The PCR fragments can be generated by Intact Genomics high fidelity Pfu DNA polymerase or other high-fidelity DNA polymerases, with primers having 15 to 18 bases of homology at their linear ends to where the product need to fuse. The linearized vector can be generated by PCR or restriction enzymes. The kit is so robust that multiple DNA fragments can be assembled simultaneously and cloned into one construct in a single reaction step within short times (usually 10-30 min) with more than 95% cloning efficiency.

Benefits

• Clone any insert at any site within any vector
• Restriction enzyme and phosphatase free system
• Joining multiple large fragments at once
• Precise insertion at a desired orientation
• Rapid and high efficiency with > 95% positive clones

Quality Control 
Quality control is performed following the production of each new lot of product to ensure that it meets the quality standards and specifications designated for the product. Each lot is repeatedly compared side-by-side with leading competitors to ensure our products outperform the competitor before product launching. 

Contents & Storage

• 5x ig-Fusion enzyme premix: -20 °C
• 2x PCR premix: -20 °C
• High efficiency competent cells: -80 °C
• Recovery medium:4 °C or -20 °C

Protocol
1. Linearize the vector by restriction enzyme digestion or inverse PCR and purify the product with spin column.
2. Design PCR primers for the gene of interest with 15 to 20 bp at 5'-extensions that are complementary to the ends of the linearized vector.
3. Amplify the gene of interest with Intact Genomics 2x PCR premix or any other high-fidelity DNA polymerase. Run the PCR product on an agarose gel to determine the integrity of the PCR product.
4. Purify the PCR product with spin column.
5. Set up the ig-Fusion cloning reaction as follows: Insert and vector molar ratio 3:1 produce the highest number of colonies.  

6.  Mix the reaction mixture thoroughly.
7.  Incubate the reaction mixture at 50 °C for 10-30 min, then place on ice. Number of colonies depend on the incubation time, insert size and number of inserts need to clone.
8.  Use 2.0 µl of the reaction mixture and transform into high efficiency ig 10B chemical or electroporation competent cells (included). To get the maximum number of colonies, we recommend to use ig 10B electrocompetent cells (Cat # 1212).

RT 2X Master Mix

• Up to 9kb cDNA synthesis
• Ensures sample to sample consistency
• Large RNA sample volume capacity
• Ready to use

Size: 100 rxns

RT 2X Master Mix is a proprietary, ready-to-use master mix for first-strand cDNA synthesis in a 2X
concentration. This optimized reaction mix contains ribonuclease inhibitor, dNTPs, and a balanced
concentration for oligo(dT) and random primers. The ribonuclease inhibitor effectively protects RNA
template from degradation. The oligo(dT) anneals selectively to the poly(A) tail of mRNAs and the
random primers do not require the presence of poly(A) and they are utilized for the transcription of
mRNA 5-end regions. The resultant cDNA can be directly used as template in different PCR experiments.

Kit Components:

Storage: 
Store at -20°C in a frost-free freezer.

• Application:

  • cDNA synthesis
  • Construction of cDNA libraries
  • Generation of probes for hybridization

   

  Protocol:

  1. Thaw RNA templates and all reagents on ice. Mix each solution by vortexing.
  2. Assemble the following components in a tube on ice, and mix well:

  Components	Volume	Final Conc.
  Total RNA, or	Variable	1 ng - 2 µg/rxn
  mRNA	Variable	1 pg - 2 ng/rxn
  2X Reaction Mix	10 µl	1X
  H2O	Up to 19 µl	-

  3. Heat the mixture at 65°C for 5 mins and incubate on ice for at least 1 min.
  4. Collect all components by a brief centrifugation and add 1 µl of the EasyScript RTase to the tube.
  5. Mix well and collect all the components by a brief centrifugation.
  6. Incubate the tube at room temperature for 10 min for annealing.
  7. Perform cDNA synthesis by incubating the tube for 50 min at 42°C.
  8. Stop the reaction by heating it at 85°C for 5 min.
  9. Chill on ice. The newly synthesized first-strand cDNA is ready for immediate downstream applications.