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HT ExoSAP-IT™ Fast High-Throughput PCR Product Cleanup Applied Biosystems™

HT ExoSAP-IT Fast High-Throughput PCR Product Cleanup, with its shortened 14 minute protocol, is a highly stable, customized formulation of the original ExoSAP-IT reagent. It is ideal for high-throughput PCR product cleanup using liquid handling platforms and multi-channel pipettes. Multiple packaging formats (vials, 8-tube strips, 96-well plates) ensure ease of use and flexibility in experimental design.

Like the original formulation, HT ExoSAP-IT Fast reagent is a mixture of Exonuclease I and Shrimp Alkaline Phosphatase for the enzymatic removal of excess primers and dNTPs following a PCR reaction. This enzymatic cleanup reagent has a decreased viscosity yet maintains the same convenience and stability that you have come to expect from the ExoSAP-IT product line. As with all products in the ExoSAP-IT family, HT ExoSAP-IT Fast reagent allows PCR cleanup with a single pipetting step. .

HT ExoSAP-IT Fast reagent is scalable for PCR cleanup from a single tube up to a 384-well plate PCR reaction.
•14 minute protocol is half the time of standard protocols
•One-tube PCR cleanup — add HT ExoSAP-IT Fast reagent directly to PCR product
•100% sample recovery — no loss of PCR products regardless of the fragment size
•Stable at 4°C for one month
•Removes excess primers and dNTPs — does not interfere with downstream applications
   -sequencing
   -SNP analysis
   -single base extension
   -fragment analysis

Adding to our family of patented ExoSAP-IT products, HT ExoSAP-IT Fast High-Throughput PCR Cleanup reagent quickly provides accurate and consistent results in high-throughput applications. HT ExoSAP-IT Fast reagent is over 50% faster than standard HT ExoSAP-IT reagent with a 14 minute total protocol time.
•Add it directly to the PCR product and incubate at 37°C for just 7 minutes.
• After PCR treatment, inactivate it by simply heating to 80°C for another 7 minutes.

This product is ideal for high volume labs that require quick sample turn around. With such a brief and simple protocol, proper sample cleanup never needs to be sacrificed due to processing or instrument time constraints. Ensure the highest quality Sanger sequencing results while keeping sample processing moving quickly and efficiently.

PCR products cleaned up with HT ExoSAP-IT Fast reagent prior to automated sequencing displayed superior sequencing results compared to the untreated equivalent (Fig. 2).

HT ExoSAP-IT Fast reagent displayed excellent stability in all formats tested including vials, 8-tube strips, and 96-well plates. The product showed no loss in function after 10 freeze thaw cycles (Fig. 3), and is stable at variable temperatures for extended periods of time compared to enzyme mix equivalents. With excellent stability and a greater than 50% faster protocol, HT ExoSAP-IT Fast reagent is ideal for high-throughput assays where robotic platforms are utilized and large volumes of samples are processed in successive testing runs.

Collectively, these results clearly demonstrate the effectiveness of HT ExoSAP-IT Fast reagent to ensure the highest quality sequences while keeping sample processing moving quickly and efficiently.

ExoSAP-IT™ Express PCR Product Cleanup Reagent Applied Biosystems™

Key features of ExoSAP-IT™ Express reagent:
5 minute protocol — Fastest turnaround time to results
One-tube, one-step PCR cleanup — Add ExoSAP-IT Express directly to PCR product
Novel enzyme technology — Irreversibly inactivated in just one minute at 80°C
Conserve PCR samples — 100% recovery of PCR products, regardless of amplicon length
Eliminate spin columns or beads — Significantly decreases time and errors while increasing yield
Compatible — No interference in downstream applications
Scalable — Treat a wide range of volumes
Multiple formats — Single tube, 8 tube strip, and 96-well plate formats offer flexibility from manual pipetting to automated workflows
Go green — Generate less waste with a single-tube solution to PCR cleanup

Overview
It is necessary to remove excess primers and unincorporated nucleotides from the PCR reaction prior to sequencing.

An enzymatic cleanup method offers precision and speed over technologies such as spin columns or beads. ExoSAP-IT Express reagent consists of two hydrolytic enzymes, a modified Exonuclease I and Shrimp Alkaline Phosphatase (SAP), in a specially formulated buffer. It is used to remove excess primers and dNTPs from a PCR mixture with no interference in downstream applications. This unique, highly stable, one tube solution allows for greater confidence and superior sequencing results (Fig. 1).

Innovative solution
Our scientists engineered Exonuclease I to have increased heat lability, allowing for inactivation in only 1 minute (Fig. 2). ExoSAP-IT Express reagent is added directly to the PCR product for a 4 minute cleanup at 37°C, followed by a 1 minute inactivation at 80°C, which provides accurate and consistent results in just 5 minutes.

Fastest PCR cleanup method
This novel technology allows for a significant reduction in sample cleanup time with minimal steps, providing the simplest workflow (Fig. 3). ExoSAP-IT Express reagent offers rapid turnaround times and improved efficiency of resources while delivering the quality that scientists have become accustomed to when using ExoSAP-IT for PCR product cleanup.

Conserve PCR samples — simple one step, 100% recovery
The ExoSAP-IT Express enzymatic cleanup method is designed to minimize errors by reducing your protocol to a single pipette step, allowing for automated or manual processing from a single tube or microtiter well. ExoSAP-IT Express reagent outperforms the competition by ensuring 100% recovery of all amplicon sizes, both short and long (Table 1).

ExoSAP-IT Express PCR Product Cleanup is active in commonly used PCR buffers, so no buffer exchange is required. After treatment, ExoSAP-IT Express reagent is inactivated by heating at 80°C for 1 minute. The treated PCR products are then ready for subsequent analysis in applications that require DNA to be free of excess primers and nucleotides.

Sphingosine Kinase 1 Recombinant Protein Invitrogen™

Human SPHK1 Recombinant Protein for Ctrl

HT ExoSAP-IT™ High-Throughput PCR Product Cleanup Applied Biosystems™

HT ExoSAP-IT High-Throughput PCR Product Cleanup is an alternative formulation of ExoSAP-IT reagent specifically designed for the unique requirements of high-throughput, automated platforms, and multi-channel pipettes. HT ExoSAP-IT reagent has both a longer lifetime at higher temperatures and a decreased viscosity for robotic pipetting.

Like ExoSAP-IT for PCR Product Cleanup (PN 78200), HT ExoSAP-IT reagent is a mixture of Exonuclease I and Shrimp Alkaline Phosphatase (SAP) which removes excess primers and dNTPs following a PCR reaction in a single incubation. HT ExoSAP-IT reagent possesses greater thermal stability and lower viscosity which provides greater ease-of-use in automated 96- and 384-well formats.

One-tube PCR cleanup — Add HT ExoSAP-IT reagent directly to PCR product
Exceptional accuracy — Achieve high quality data even with long read lengths
10% sample recovery — No loss of PCR products regardless of the fragment size
Simple, single-step — Replaces the multiple steps and wasted time inherent with beads or columns
High-throughput processing — Even faster time to results with a low viscosity formulation, allowing robotic pipetting
Removes excess primers and dNTPs — Does not interfere with downstream applications
  – sequencing
  – SNP analysis
  – single base extension
  – fragment analysis
  – in vitro transcription
Scalable — Treat reaction volumes from 5 micro;L to 5 L
Convenient packaging — Available in 8-tube strips and 12 strips in a 96-well plate
Stable at +25deg;C for 8 hours — Retains full functional activity and at 45°C is stable for one week

HT ExoSAP-IT reagent is added directly to the PCR product and incubated at 37°C for 15 minutes. After PCR treatment, it is inactivated simply by heating to 80°C for 15 minutes. Our high-throughput formulation is designed for robotic pipetting and multi-channel pipettes. HT ExoSAP-IT reagent has a lower viscosity than the standard mix and is available in an 8-tube strip format for ease of use.

HT ExoSAP-IT reagent utilizes two hydrolytic enzymes, Exonuclease I and SAP, together in a specially formulated buffer, to remove unwanted dNTPs and primers from PCR products. Exonuclease I removes residual single-stranded primers and any extraneous single-stranded DNA produced in the PCR. SAP removes the remaining dNTPs from the PCR mixture.

Simple: Single-step
HT ExoSAP-IT reagent requires only one pipetting step and two incubations. Just add HT ExoSAP-IT reagent to the PCR product and within 30 minutes sequencing, SNP analysis, single base extension, or fragment analysis can be performed. Enzymatic removal of excess primers and unincorporated nucleotides occurs in one easy step by using HT ExoSAP-IT reagent in a single tube, 8-tube strip, or 12 strips in a 96-well low skirted PCR plate. Our new formatting makes this an ideal product for use with robotics.

Exceptional accuracy with HT ExoSAP-IT
Achieve high data quality and sequencing accuracy with HT ExoSAP-IT, even with long read lengths. Sequence reads of PCR products treated with HT ExoSAP-IT were on average 50+ bases longer than samples treated with competitor products. Using human genomic DNA, a 1007 bp fragment was sequenced with no miscalls. The size limitations associated with alternative PCR cleanup methods are not a factor with HT ExoSAP-IT. Phred 20 values over the entire length of a 970 bp sequence was 822 ± 9 with HT ExoSAP-IT-treated samples and only 776 ± 83 with samples treated with Agencourt™ AMPure™ XP.

No sample loss
Use of HT ExoSAP-IT eliminates all gel or column purifications, sedimentations, filtrations, beads, and/or magnetic separations(1). There is 100% recovery of both short and long PCR products with HT ExoSAP-IT.

HT ExoSAP-IT-treated PCR product is stable
Ht ExoSAP-IT reagent is rigorously tested and subject to strict quality control. No product degradation occurred after storage of HT ExoSAP-IT treated PCR product for one week at 24°C.

References:
1. DUGAN, K. A., LAWRENCE, H. S., HARES, D. R., FISHER, C. L. AND BUDOWLE B. (2002) J. Forensic Sci 47, 811-818.
2. HANKE, M. AND WINK, M. (1994) BioTechniques 17, 858-860.
3. MU, J., DUAN, J., MAKOVA, K., JOY, D., HUYNH, C., BRANCH, O., LI, W. AND SU, X. (2002) Nature 418, 323-326.
4. SILVA, JR., W. A., COSTA, M. C. R., VALENTE, V., DE FREITAS SOUSA, J., PACÓ-LARSON, M. L., ESPREAFICO, E. M., CAMARGO, S. S., MONTEIRO, E., DE JESUS, A., HOLANDA, M. A., ZAGO, M. A., SIMPSON, A. J. G. AND NETO, E. D. (2001) BioTechniques 30, 537-542.
5. WERLE, E., SCNEIDER C., RENNER, M., VÖLKER, M. AND FIEHN, W. (1994) Nucleic Acids Res. 22, 4354-4355.

PCR Product Pre-Sequencing Kit Applied Biosystems™

The PCR Product Pre-Sequencing Kit uses a novel, enzymatic clean-up method to pre-treat PCR products prior to sequencing, without any subsequent purification or separation steps. The two hydrolytic enzymes used in this kit, shrimp alkaline phosphatase and exonuclease I, effectively remove excess dNTPs and primers present in the final PCR product reaction mixture. The enzymes are conveniently added to an aliquot of the PCR product mixture, incubated at 37°C, and then inactivated at 80°C. The result is a cleaner PCR product, free from excess nucleotides and primers, and ready to be sequenced with standard sequencing reagents.

Human MMP3 Recombinant Protein Invitrogen™

Human MMP3 Recombinant Protein for Ctrl

Sphingosine Kinase 2 Recombinant Protein Invitrogen™

Human SPHK2 Recombinant Protein for Ctrl

Sphingosine Kinase 1a Recombinant Protein Invitrogen™

Mouse SPHK1a Recombinant Protein for Ctrl

Human ADAMTS5 Recombinant Protein Invitrogen™

Human ADAMTS5 Recombinant Protein for Ctrl

Human MMP2 Recombinant Protein Invitrogen™

Human MMP2 Recombinant Protein for Ctrl

TrueCut™ Cas9 Protein v2 Invitrogen™

Invitrogen TrueCut Cas9 Protein v2 is a next-generation CRISPR Cas9 protein engineered to deliver maximum editing efficiency. Features include:

• Consistently high editing efficiency in all tested cell lines including standard, immune, primary, and stem, with up to 2X higher editing efficiency in difficult targets than the competition
• High quality—manufactured under strict ISO 13485 quality standards
• Validated protocols for a large number of cell types help you achieve success faster

TrueCut Cas9 Protein v2 is recombinant Streptococcus pyogenes Cas9 (wt) protein, purified from E. coli, for genome editing with CRISPR technology. Cas9 protein forms a very stable ribonucleoprotein (RNP) complex with the guide RNA (gRNA) component of the CRISPR/Cas9 system. Incorporation of nuclear localization signals (NLS) aids delivery to the nucleus, increasing the rate of genomic DNA cleavage.

• Transfection-ready, using either lipid-mediated transfection reagents or electroporation
• Eliminates time-consuming cloning steps
• Available in 2 concentrations:
    --1 μg/μL for use in standard editing scenarios
    --5 μg/μL for optimization of editing conditions in more difficult scenarios such as in primary or embryonic cells, microinjection, or when screening multiple gRNA sequences at a time
    --For custom sizes or concentrations, please inquire

Options for obtaining CRISPR gRNA for use with TrueCut Cas9 Protein v2:
1. Order transfection-ready TrueGuide synthetic gRNAs
2. Generate transfection-ready gRNA in as little as four hours including template assembly with our GeneArt Precision gRNA Synthesis Kit
3. Have our Custom Services team design, synthesize, and purify in vitro (IVT) gRNA sequences for you

HT ExoSAP-IT™ Fast High-Throughput PCR Product Cleanup Applied Biosystems™

HT ExoSAP-IT Fast High-Throughput PCR Product Cleanup, with its shortened 14 minute protocol, is a highly stable, customized formulation of the original ExoSAP-IT reagent. It is ideal for high-throughput PCR product cleanup using liquid handling platforms and multi-channel pipettes. Multiple packaging formats (vials, 8-tube strips, 96-well plates) ensure ease of use and flexibility in experimental design.

Like the original formulation, HT ExoSAP-IT Fast reagent is a mixture of Exonuclease I and Shrimp Alkaline Phosphatase for the enzymatic removal of excess primers and dNTPs following a PCR reaction. This enzymatic cleanup reagent has a decreased viscosity yet maintains the same convenience and stability that you have come to expect from the ExoSAP-IT product line. As with all products in the ExoSAP-IT family, HT ExoSAP-IT Fast reagent allows PCR cleanup with a single pipetting step. .

HT ExoSAP-IT Fast reagent is scalable for PCR cleanup from a single tube up to a 384-well plate PCR reaction.
•14 minute protocol is half the time of standard protocols
•One-tube PCR cleanup — add HT ExoSAP-IT Fast reagent directly to PCR product
•100% sample recovery — no loss of PCR products regardless of the fragment size
•Stable at 4°C for one month
•Removes excess primers and dNTPs — does not interfere with downstream applications
   -sequencing
   -SNP analysis
   -single base extension
   -fragment analysis

Adding to our family of patented ExoSAP-IT products, HT ExoSAP-IT Fast High-Throughput PCR Cleanup reagent quickly provides accurate and consistent results in high-throughput applications. HT ExoSAP-IT Fast reagent is over 50% faster than standard HT ExoSAP-IT reagent with a 14 minute total protocol time.
•Add it directly to the PCR product and incubate at 37°C for just 7 minutes.
• After PCR treatment, inactivate it by simply heating to 80°C for another 7 minutes.

This product is ideal for high volume labs that require quick sample turn around. With such a brief and simple protocol, proper sample cleanup never needs to be sacrificed due to processing or instrument time constraints. Ensure the highest quality Sanger sequencing results while keeping sample processing moving quickly and efficiently.

PCR products cleaned up with HT ExoSAP-IT Fast reagent prior to automated sequencing displayed superior sequencing results compared to the untreated equivalent (Fig. 2).

HT ExoSAP-IT Fast reagent displayed excellent stability in all formats tested including vials, 8-tube strips, and 96-well plates. The product showed no loss in function after 10 freeze thaw cycles (Fig. 3), and is stable at variable temperatures for extended periods of time compared to enzyme mix equivalents. With excellent stability and a greater than 50% faster protocol, HT ExoSAP-IT Fast reagent is ideal for high-throughput assays where robotic platforms are utilized and large volumes of samples are processed in successive testing runs.

Collectively, these results clearly demonstrate the effectiveness of HT ExoSAP-IT Fast reagent to ensure the highest quality sequences while keeping sample processing moving quickly and efficiently.

ExoSAP-IT™ PCR Product Cleanup Applied Biosystems™

Features and Benefits of the ExoSAP-IT™ PCR Product Cleanup
Conserves PCR samples — 100% recovery of both short and long PCR products
One tube/one step PCR cleanup — Add ExoSAP-IT reagent directly to PCR product
Eliminates spin columns — Decreases time and expense while increasing yield
Removes contaminating primers and dNTPs — No interference in downstream applications
Scalable — Economical for high-throughput purification
Simple processing — Robotic-friendly; Replaces beads, filtrations, and plates
Generates less waste than columns

ExoSAP-IT reagent is designed for simple, quick PCR cleanup for downstream applications, such as DNA sequencing or Single Nucleotide Polymorphism (SNP) analysis. When PCR amplification is complete, any unconsumed dNTPs and primers remaining in the PCR product mixture will interfere with these methods. ExoSAP-IT removes these contaminants.

ExoSAP-IT is added directly to the PCR product and incubated at 37°C for 15 minutes (Fig. 1). After PCR treatment, ExoSAP-IT is inactivated simply by heating to 80°C for 15 minutes.

ExoSAP-IT single-step PCR cleanup utilizes two hydrolytic enzymes, Exonuclease I and Shrimp Alkaline Phosphatase (SAP), together in a specially formulated buffer, to remove unwanted dNTPs and primers from PCR products. Exonuclease I removes residual single-stranded primers and any extraneous single-stranded DNA produced in the PCR. SAP removes the remaining dNTPs from the PCR mixture.

Rapid PCR Product Cleanup Protocol
ExoSAP-IT requires only one pipetting step and two incubations. Just add ExoSAP-IT to the PCR product and within 30 minutes sequencing or SNP analysis can be performed.

Simple: Single-Step
The method is designed to require a minimum of ‘hands-on’ time. Enzymatic removal of excess primers and unincorporated nucleotides occurs in one easy step by using ExoSAP-IT reagent in a single tube or microtiter well. Only simple pipette transfers are required, therefore, many samples can be processed at once, either manually or with robotics.

No Sample Loss
Use of ExoSAP-IT reagent eliminates all gel or column purifications, sedimentations, filtrations, beads, and/or magnetic separations. There is 100% recovery of both short and long PCR products with ExoSAP-IT (Fig. 2).

Achieve High Data Quality from PCR Products
ExoSAP-IT reagent may be used as an effective cleanup method prior to fluorescent or radioactive DNA sequencing, SNP analysis, or any other application requiring a PCR product free of excess nucleotides and primers.

References:
DUGAN, K. A., LAWRENCE, H. S., HARES, D. R., FISHER, C. L. AND BUDOWLE B. (2002) J. Forensic Sci 47, 811-818.
HANKE, M. AND WINK, M. (1994) BioTechniques 17, 858-860.

MU, J., DUAN, J., MAKOVA, K., JOY, D., HUYNH, C., BRANCH, O., LI, W. AND SU, X. (2002) Nature 418, 323-326.
SILVA, JR., W. A., COSTA, M. C. R., VALENTE, V., DE FREITAS SOUSA, J., PACÓ-LARSON, M. L., ESPREAFICO, E. M., CAMARGO, S. S., MONTEIRO, E., DE JESUS, A., HOLANDA, M. A., ZAGO, M. A., SIMPSON, A. J. G. AND NETO, E. D. (2001) BioTechniques 30, 537-542.
WERLE, E., SCNEIDER C., RENNER, M., VÖLKER, M. AND FIEHN, W. (1994) Nucleic Acids Res. 22, 4354-4355.

FHIT Recombinant Protein Invitrogen™

Human FHIT Recombinant Protein for Ctrl

ExoSAP-IT Express PCR Product Cleanup with Tracking Dye Applied Biosystems™

Our fastest PCR cleanup method - just 5 minutes to superior sequencing results. Now available with optional tracking dye.

ExoSAP-IT™ Express reagent:
5 minute protocol — Fastest turnaround time to results
One-tube, one-step PCR cleanup — Add ExoSAP-IT Express directly to PCR product
Novel enzyme technology — Irreversibly inactivated in just one minute at 80°C
Conserve PCR samples — 100% recovery of PCR products, regardless of amplicon length
Eliminate spin columns or beads — Significantly decreases time and errors while increasing yield
Compatible — No interference in downstream applications
Scalable — Treat a wide range of volumes
Multiple formats — Single tube, 8 tube strip, and 96-well plate formats offer flexibility from manual pipetting to automated workflows
Go green — Generate less waste with a single-tube solution to PCR cleanup
Optional tracking dye — Never lose track of reagent addition during pipetting

Overview
It is necessary to remove excess primers and unincorporated nucleotides from the PCR reaction prior to sequencing. An enzymatic cleanup method offers precision and speed over technologies such as spin columns or beads. ExoSAP-IT Express reagent consists of two hydrolytic enzymes, a modified Exonuclease I and Shrimp Alkaline Phosphatase (SAP), in a specially formulated buffer. It is used to remove excess primers and dNTPs from a PCR mixture with no interference in downstream applications. This unique, highly stable, one tube solution allows for greater confidence and superior sequencing results (Fig. 1).

Innovative solution
Affymetrix scientists engineered Exonuclease I to have increased heat lability, allowing for inactivation in only 1 minute (Fig. 2). ExoSAP-IT Express reagent is added directly to the PCR product for a 4 minute cleanup at 37°C, followed by a 1 minute inactivation at 80°C, which provides accurate and consistent results in just 5 minutes.

Fastest PCR cleanup method
This novel technology allows for a significant reduction in sample cleanup time with minimal steps, providing the simplest workflow (Fig. 3). ExoSAP-IT Express reagent offers rapid turnaround times and improved efficiency of resources while delivering the quality that scientists have become accustomed to when using ExoSAP-IT for PCR product cleanup.

Optional tracking dye
ExoSAP-IT Express reagent is also available premixed with a tracking dye for added convenience. This inert, blue color dye
offers visual confirmation that the reagent has been added to samples for PCR cleanup during the pipetting process. The dye does not interfere with downstream applications such as subsequent PCR and sequencing.

Conserve PCR samples — simple one step, 100% recovery
The ExoSAP-IT Express enzymatic cleanup method is designed to minimize errors by reducing your protocol to a single pipette step, allowing for automated or manual processing from a single tube or microtiter well. ExoSAP-IT Express reagent outperforms the competition by ensuring 100% recovery of all amplicon sizes, both short and long (Table 1).

ExoSAP-IT Express PCR Product Cleanup is active in commonly used PCR buffers, so no buffer exchange is required. After treatment, ExoSAP-IT Express reagent is inactivated by heating at 80°C for 1 minute. The treated PCR products are then ready for subsequent analysis in applications that require DNA to be free of excess primers and nucleotides.

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