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PRODUCT

Research sgRNA

Early discovery breakthroughs in therapeutic development begin with research single guide RNA (sgRNA).

Overview

Synthego's Research sgRNA Overview

  • Yields: 1.5, 3, 5, 10, 20, 30, 50, 100, >100* nmol
  • Length: 20-115 nucleotides, >115 nucleotides*
  • Modifications: Standard modification, custom modifications*, and Unmodified
  • Purification: solid phase extraction (standard), HPLC* for yields greater than 100 nmol

We made it easy to order sgRNA with our user-friendly order interface.

*For gRNA lengths >115 nucleotides, HPLC purification, and/or larger yield options, schedule a call with our sales team and select 'Synthetic sgRNA' as your ‘Product of Interest’ on the form. Our team is excited to discuss how we can support your projects.

Reasons to use our Research sgRNA

Compatible With Your Nuclease: For CRISPR experiments using novel, engineered, or well-established nucleases, our Research sgRNA are best-in-class enabling you to maximize your nucleases' efficiency for early therapeutic development and discovery research.

Assess More Modifications For Precision: Our standard modifications, which include 2'-O-methyl (OMe) analogs and 3' phosphorothioate (PS) linkages, enhance gRNA stability and performance. For nuclease testing requiring custom placement of the OMe analog and PS linkages, our Research sgRNA allows you to precisely select their locations to optimize your gRNA for your experiments.

Explore Alternative Modifications: CRISPR experimentation is diverse, and OMe analogs and PS bonds are just the beginning. If your CRISPR experiments require alternative sgRNA chemical modifications beyond OMe analogs and PS bonds, our Research sgRNA are fully customizable to support your unique research goals.

Designing Your eSpOT-ON, hfCas12Max, AccuBase™, and SpCas9 Guide RNA

Try hfCas12Max in your workflow with our validated gRNAs.
Gene Name hfCas12Max Target Sequence
B2M TATCTCTTGTACTACACTGA
TRAC GAGTCTCTCAGCTGGTACAC

*Validated in human cell lines (HEK293T, T Cells, and iPSC). hfCas12Max gRNAs are not compatible with eSpOT-ON or SpCas9. SpCas9 sgRNA controls can be purchased here. eSpOT-ON and AccuBase controls can be purchased through our sales team.

To design your own guide RNA, select your target sequences using your favorite CRISPR design platform, such as CHOPCHOP, Benchling, CRISPOR, or Cas-Designer CRISPR RGEN Tools. PAM sequences for eSpOT-ON, hfCas12Max, and SpCas9 are below:

  • eSpOT-ON has a canonical PAM 5' - NGG - 3'
  • hfCas12Max has a broadened PAM 5'-TN-3' or 5'-TTN-3'
  • AccuBase has a canonical PAM 5' - NGG - 3'
  • spCas9 has a canonical PAM 5' - NGG - 3'

To order your guide RNA, upload your target sequences without the PAM sequence into our easy-to-use ordering interface so we can synthesize your guide RNA.

Synthego's standard chemical modifications

Incorporating our standard chemical modifications into our gene editors' gRNA enhances their performance. The table below outlines the modifications used with each editor.
CRISPR gene editor Synthego's standard chemical modification pattern
eSpOT-ON recombinant protein
eSpOT-ON mRNA
2'-O-Methyl analog at the first 3 and last 3 bases and 3' phosphorothioate bonds between 3 first and last 4 bases
hfCas12Max recombinant protein 2'-O-Methyl analog at the first 3 bases. The last 4 bases have 3 modifications ending with a nonmodified base. With 3' phosphorothioate bonds between 3 first and last 4 bases.
AccuBase cytosine base editor protein 2'-O-Methyl at 3 first and last bases, 3' phosphorothioate bonds between first 3 and last 2 bases
SpCas9 recombinant protein 2'-O-Methyl at 3 first and last bases, 3' phosphorothioate bonds between first 3 and last 2 bases

hfCas12Max paired with best-in-class Research sgRNA

Complexing hfCas12Max nuclease with our best-in-class Research sgRNA achieves consistently high editing efficiencies. If you are seeking alternatives CRISPR gene editing methods, hfCas12Max is the nuclease you need for your next CRISPR experiment or therapeutic development.

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Different hfCas12Max Nuclease:sgRNA ratios were tested to evaluate editing efficiency in HEK293T cells. It is clear when hfCas12Max is complex with our Research sgRNA, it can achieve exceptionally high editing efficiencies.

Learn More about our high-fidelity Cas12 nuclease, hfCas12Max, as it sets the stage for being the next alternative nuclease used to develop life-saving cell and gene therapies using CRISPR.

SpCas9 and our Research sgRNA lead to unmatched results

SpCas9 nuclease is widely used to perform CRISPR experiments. When complexed with our best-in-class Research sgRNA, SpCas9 achieves editing efficiencies that are unattainable with other sgRNA options. High-efficiency CRISPR T cell editing can be achieved to develop CAR-T cell therapies and fight viral infections.

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Synthego’s Research sgRNAs demonstrate superior editing efficiency compared to other vendors at the CD46 loci in resting human CD4+ T Cells. Synthego Research sgRNAs were compared against another vendor’s sgRNA and 2-part (crRNA:trRNA), demonstrating consistently high (60%+) knockout efficiency regardless of preparation conditions.

Best-in-class Research sgRNA that overcome editing challenges

Not all cell types are easy to perform CRISPR experiments in. Certain cell types, like human resting T cells, can be challenging to achieve high editing efficiencies while maintaining high cell viability. Our best-in-class Research sgRNA and SpCas9 nuclease can overcome these challenges in even the most difficult cell types.

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High editing efficiency and viability were observed in activated human primary T cells. Robust gene knockout is demonstrated in human primary T cell knockout using Synthego’s Research sgRNA complexed with its SpCas9 nuclease and transfected via nucleofection. High editing efficiency (>70%) of various loci was observed in activated human primary CD4+/CD8+ T cells across two independent donors (HD-A, HD-B) as assessed by sequencing analysis. Additionally, high cell viability (>95%) was consistently achieved, irrespective of editing locus, as assessed by flow cytometry.

CRISPR Nuclease Comparison of SpCas9 vs hfCas12Max

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RNP complex schematics of eSpOT-ON (left), SpCas9 (center), and hfCas12Max (right). Depicted in the schematic are genomic DNA (light blue), PAM sequence (pink), and predicted cut site locations on genomic DNA (gray scissors). The guide RNA (gRNA) is comprised of the target sequence (green) and scaffold (dark blue).

eSpOT-ON vs SpCas9 vs hfCas12Max

eSpOT-ON Nuclease (protein or mRNA)

SpCas9 Nuclease

hfCas12Max Nuclease

Description Engineered from PsCas9 to achieve high on-target editing and minimized off-target editing The nuclease that led to Jennifer Doudna and Emmanuelle Charpentier's Nobel Prize. Engineered from Cas12i to achieve higher on-target editing and minimized off-target editing.
Size 1409 amino acids 1368 amino acids 1080 amino acids
PAM Sequence (N = any nucleotide)

5'-NGG-3'PAM

PAM 3' of the target DNA sequence

5'-NGG-3’

PAM is 3’ of the target DNA sequence

5'-TN-3' or 5'-TTN-3'
DNA Cleavage

Staggered-cut

cleavage on the target strand occurs 3nt upstream of the PAM, while the non-target strand is cut 6-7nt upstream of the PAM.

Blunt end cut

Cleavage 3nt upstream of PAM sequence

Staggered-cut

Cleavage on the target strand occurs 24nt downstream from PAM, while the non-targeted strand is cut 14-16nt downstream.

Endonuclease Domains HNH and RuvC HNH and RuvC RuvC
gRNA Length and Target Sequence

109nt

22nt target sequence

97 - 103nt

20nt Target Sequence

44 - 50nt

20nt Target Sequence

gRNA Components crRNA + tracrRNA as a single guide RNA (sgRNA) crRNA + tracrRNA as a single guide RNA (sgRNA) crRNA
Variants or Origin Engineered from Parasutterella secunda (PsCas9)

Type IIa CRISPR-Cas system of Streptococcus pyogenes Cas9

Type V CRISPR-Cas system of Cas12

Are you ready to make a seamless transition from early discovery to clinical trials?

Our continuum of CRISPR solutions enables therapeutic developers to transition seamlessly from early discovery breakthroughs to clinical trials - research to GMP. With over +1 million sgRNA synthesized and used across various applications, researchers know the benefits of our best-in-class sgRNA because they provide them the flexibility in scale, length, and purity to suit their specific needs.

Within our continuum of CRISPR solutions, our Research sgRNA are sought out by scientists to start their therapeutic development because they can achieve editing efficiencies up to 99% and can easily scale during the development phase of their work. Synthesized for early discovery and research applications, breakthroughs in therapeutics and science begin with our best-in-class Research sgRNA.

When transitioning from discovery research to preclinical studies and clinical applications, it’s crucial to use high-quality sgRNA from a trusted manufacturer with a proven track record in synthesizing GMP-grade sgRNA that meets current good manufacturing practice standards. However, not all GMP materials adhere to these standards. Synthego has been producing GMP sgRNA that meets regulatory requirements for years, helping scientists seamlessly transition their therapeutic development through each phase toward clinical application. If you're preparing to move your therapeutic into preclinical studies or clinical trials, contact us to learn how we can support your journey.

Synthego really stands out because I'm not sure if there are many others that can offer screening scale, research use guides for your lead discovery, medium to large scale RUO guides for your preclinical and pharmacology studies, and ultimately larger scale GLP Tox and clinical grade material for early and hopefully ultimately late phase trials.
Mark DeWitt, Ph.D.
ASSOCIATE DIRECTOR, MAMMOTH BIOSCIENCES
SGRNA PURITY

Exceptional editing efficiencies without requiring HPLC purification

Not all chemically synthesized sgRNA are manufactured equally. During synthesis, some oligomers fail to extend during each cycle resulting in undesired truncated molecules mixed in with your desired sgRNA. Additionally, several types of chemical contaminants can be introduced through the manufacturing process of sgRNA. The presence of both these impurities can increase off-target binding, reduce on-target binding, or both. Several vendors offer high-performance liquid chromatography (HPLC) as a purification step to eliminate the unwanted truncated oligomers (n-3 to n-5) and contaminants. However, each vendor might have different synthesis and quality standards to reach the purity levels desired for your sgRNA.

Synthego’s best-in-class sgRNA can achieve exceptional editing efficiencies through standard manufacturing processes (non-HPLC) compared to HPLC purified sgRNA from other vendors. If the purity was substantially better, you would expect the HPLC purified sgRNA to perform better in all cases. However, that is not always the case in our study.

Depending on your application, particularly sensitive applications like therapeutic development require sgRNA that achieves high performance in early development and highly pure sgRNA as development progresses towards the clinic. To learn more about our sgRNA purity and which of Synthego’s best-in-class sgRNA is right for your application, contact our sales team here.

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Dose-dependent editing efficiency (mean ± SD) of Vendor T HPLC purified sgRNA (black), Synthego non-HPLC purified sgRNA (green), and Synthego HPLC purified sgRNA (blue) for gene targets a) RELA and b) CDC42BPB. HEK293 cells were transfected with RNPs of increasing molar ratio (SpCas9 constant at 10 μM) using nucleofection. Indel frequencies were analyzed via Sanger sequencing methods.
DELIVERABLES

Order Research sgRNA that fit your early discovery and research needs

Synthego’s Research sgRNA are explicitly designed for research purposes only and should not be used for human application, including but not limited to clinical trials and diagnostics. If you are seeking higher purity sgRNA or require more in-depth Quality Control documentation not provided by our research sgRNA deliverables (see table below), contact our Sales Team for further information.

Deliverables
  • Synthetic Research sgRNA in tube or plate format (dry)
  • One 1.5 mL Nuclease Free TE Buffer (10 mM Tris, 1 mM EDTA - ph 8.0) - for every 50 nmol of gRNA and/ or sgRNA
  • One 1.5mL Nuclease Free Water - for every 50 nmol of gRNA and/ or sgRNA
gRNA Length hfCas12Max (PAM sequence 5’-TN or 5’-(T)TNN)
  • 5' - hfCas12Max Scaffold + [17 - 23bp] - 3'
SpCas9 (PAM Sequence 5' - NGG)
  • 5' - [17 - 23bp] + SpCas9 Scaffold - 3'
Custom gRNA
  • Full length gRNA sequence required. Full length gRNA sequence includes target sequence and scaffold.

Note: The PAM sequence is not included in the final guide RNA design.

Modification hfCas12Max
  • Modified (2'-O-Methyl analog at first 3 and last 4 bases, 3' phosphorothioate bonds between 3 first and last bases)
Spcas9
  • unmodified
  • Modified (2'-O-Methyl analog first and last 3 bases; 3' phosphorothioate between first 3 and last 2 bases)
Custom gRNA
  • unmodified
  • Modified (2'-O-Methyl analog first and last 3 bases; 3' phosphorothioate between first 3 and last 2 bases)
  • custom chemical modifications*
Yield Offerings

Tube (nmol): 1.5, 3, 5, 10, 20, 30, 50, 100

Tube (nmol): >100*

Plate (nmol): 1.5, 3, 5, 10

QC and Purification
  • Standard (Electrospray Ionization Mass Spectrometry)
  • High-Performance Liquid Chromatography (HPLC)*