CRISPR Synthetic sgRNA

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Overview

CRISPR/Cas9 technology is a powerful tool for creating gene knock-ins and knock-outs. Using the ribonucleoprotein (RNP) system for delivering CRISPR/Cas9 has many advantages over the traditional plasmids or virus based delivery methods.

Advantages

DNA free

Detectable at high levels shortly after transfection

Quickly cleared from the cell for less off-target effects

Highly efficient even in hard-to-transfect cells

Best for in vivo studies

Related Services

GenScript provides comprehensive gRNA formats to support your CRISPR needs:

EasyEdit sgRNA

The easy yet effective one-step solution

Synthetic single guide RNAs (sgRNAs) with modifications supporting easy and effective editing

✔ Option to deliver in 96-well plates supporting high throughput screening

Ready to use sgRNA

No in vitro transcription or annealing
step needed

High KO efficiency

Even in hard-to-transfect cells

Flexible at scale

1 to 1000+ sgRNAs 2 to 100+ nmol

Design Custom Guides Order Your Own Design Request Now

Why choose EasyEdit sgRNA? »

Why waste time on reagent preparation? Use one piece sgRNA avoiding unstable annealing

Synthetic sgRNA provides much better editing efficiency compared to IVT sgRNA
Did we mention that preparing IVT sgRNA from plasmid template takes ~4 hours in-house?

Pricing »

Product	Quantity	Pricing	Quote
EasyEdit sgRNA	2 nmol	$99	Quote
	4 nmol	$149
	10 nmol	$259
	50 nmol	$799
	100 nmol	$1,199

Add-On Items

Product	Quantity	Pricing	Quote
Custom Primer for Assessing Editing Efficiency	2 nmol	$1.2/nt	Quote
EasyEdit Human HPRT Positive Control sgRNA	2 nmol	$39	Quote
SafeEdit Human HPRT Positive Control sgRNA	2 nmol	$69	Quote
Human HPRT Primer Mix for Assessing Editing Efficiency	2 nmol	$10	Quote
GenCRISPR™ Ultra NLS-Cas9 Nuclease (10 mg/ml)	500 μg	$695	Quote
GenCRISPR™ Ultra NLS-Cas9 Nuclease (10 mg/ml)	1 mg	$1181.50	Quote
GenCRISPR™ Ultra eSpCas9-2NLS Nuclease (10 mg/ml)	500 μg	$1245	Quote
GenCRISPR™ Ultra eSpCas9-2NLS Nuclease (10 mg/ml)	1 mg	$2116.50	Quote
Long Single-Stranded DNA Synthesis for Gene Knock-In (151-5000 nt)	3 μg and up	Starting at $0.8/nt	Quote

SafeEdit sgRNA

Safeguard your cells

HPLC purified high purity sgRNAs with modifications ensuring minimum off-target and cytotoxicity

More than 90% purity guaranteed
Ideal for primary cells and stem cell

Minimal impact for
cell viability

Minimized off-target from truncated

One stop shop from CRISRP machinery
to knock-in templates

Design Custom Guides Order Your Own Design Request Now

Why choose SafeEdit sgRNA? »

High purity SafeEdit sgRNA ensures minimal impact for cell viability

GenScript provides add-on QC options to ensure sustained quality of your cells:

Mycoplasma testing (qPCR)
Endotoxin testing (Kinetic LAL)
Stability testing
Cytotoxicity testing (co-culture with client specified cell line)

Choose HPLC purified sgRNA to avoid off-target from truncated oligos in samples

Pricing »

Product	Quantity	Pricing	Quote
SafeEdit sgRNA	2 nmol	$199	Quote
	4 nmol	$319
	10 nmol	$579
	50 nmol	$1,699
	100 nmol	$2,499

Add-On Items

Product	Quantity	Pricing	Quote
Custom Primer for Assessing Editing Efficiency	2 nmol	$1.2/nt	Quote
EasyEdit Human HPRT Positive Control sgRNA	2 nmol	$39	Quote
SafeEdit Human HPRT Positive Control sgRNA	2 nmol	$69	Quote
Human HPRT Primer Mix for Assessing Editing Efficiency	2 nmol	$10	Quote
GenCRISPR™ Ultra NLS-Cas9 Nuclease (10 mg/ml)	500 μg	$695	Quote
GenCRISPR™ Ultra NLS-Cas9 Nuclease (10 mg/ml)	1 mg	$1181.50	Quote
GenCRISPR™ Ultra eSpCas9-2NLS Nuclease (10 mg/ml)	500 μg	$1245	Quote
GenCRISPR™ Ultra eSpCas9-2NLS Nuclease (10 mg/ml)	1 mg	$2116.50	Quote
Long Single-Stranded DNA Synthesis for Gene Knock-In (151-5000 nt)	3 μg and up	Starting at $0.8/nt	Quote

GMP Grade sgRNA

GenScript now offers GMP sgRNA- Dedicated for IND and clinical phase I material production:

State-of-the-art production facility
Clean suite with class A isolator in a class C background

Comprehensive QA/QC
Proprietary NGS method to verify sgRNA identity

Trusted by scientific partners globally
Successfully delivered 30+ GMP grade batches

GMP sgRNA Production Workflow:

Contact [email protected] to learn more or Get a Quote

Resources

Why Choose CRISPR/Cas9 Ribonucleoprotein (RNP) System
CRISPR/Cas9 system can be delivered into cells in many forms, including plasmids, lentivirus, adeno-associated virus, and ribonucleoprotein (RNP) complexes. CRISPR/Cas9 ribonucleoprotein (RNP) system are composed of Cas9 protein and either a single guide RNA (sgRNA) or a CRISPR RNA (crRNA) : trans-activating crRNA (tracrRNA) duplex.
- The crRNAs are 20 nt RNA oligonucleotides designed to be complementary to a genomic target. crRNA sequence design utilizes standard gRNA design principles and sequences can be easily selected from the GenScript gRNA database or customized using the GenScript gRNA design tool.
- The tracrRNAs are 67 nt RNA oligonucleotides which together with the crRNA and Cas9 nuclease, form the activated CRISPR/Cas9 RNP complex.
- A sgRNA is a single RNA oligo that contains both the crRNA sequence, which can identify target genomic sequence, and the tracrRNA sequence, which can form complex with Cas9 protein.
When either a sgRNA or a crRNA : tracrRNA duplex forms complex with Cas9 protein, they can together, create double strand breaks at a locus complementary to the 20 nt guide RNA sequence, 3-4 bp upstream from a protospacer adjacent motif (PAM) sequence (5'-NGG-3').

Unlike the traditional plasmids or lentivirus delivery methods, CRISPR/Cas9 RNP are delivered as intact complexes, and do not require cellular expression, thus has many advantages.
- DNA free
- Detectable at high levels shortly after transfection
- Quickly cleared from the cell for less off-target effects
- Highly efficient even in hard-to-transfect cells
- Best for in vivo studies
Recently, several studies have showed that sgRNA has better stability than crRNA:tracrRNA when duplexed with Cas9, thus leading to higher editing efficiency. Synthesis of 100 nt long sgRNAs was traditionally possible through in vitro-transcription (IVT) using phage RNA polymerase. These in vitro transcribed sgRNAs contain a 5’-triphosphate, which was thought to trigger immune response in many cell types. A recent study showed that sgRNAs with 5’-triphosphate modifications produced through in vitro-transcription can indeed induce innate immune responses and lead to cytotoxicity in human and murine cells. However, chemically synthesized sgRNAs without the 5’-triphosphate modifications demonstrated much better editing efficiency in cells, thus supporting that chemically synthesized sgRNAs are the most ideal reagent for CRISPR genome editing up till now¹.

Kim et al. CRISPR RNAs trigger innate immune responses in human cells. Genome Res. 2018. 28: 367-373.
CRISPR Handbooks and Protocols

CRISPR Handbook
Enabling Genome Editing and Transforming Life Science Research
Free Download

CRISPR/Cas9 gRNA Vector CRISPR/Cas9 gRNA Vector
Design Guide
Free Download Free Download

CRISPR RNA FAQs

How many sgRNA or crRNA sequences are needed for targeted knock-out?

A minimum of 3 crRNA sequences are recommended to ensure knock-out and experimental accuracy. Independently obtained knock-out mutants provide redundancy to safeguard against any hidden off-target effects.
What are the advantages of using synthetic sgRNA for gene editing compared to the traditional plasmids or virus based gRNA delivery methods?
Unlike the traditional plasmids or lentivirus delivery methods, CRISPR/Cas9 RNP are delivered as intact complexes, and do not require cellular expression, thus has many advantages.
- DNA free
- Detectable at high levels shortly after transfection
- Quickly cleared from the cell for less off-target effects
- Highly efficient even in hard-to-transfect cells
- Best for in vivo studies
What is the different between using synthetic sgRNA vs using crRNA: tracrRNA duplex?

When using sgRNA, there is no need for annealing crRNA and tracrRNA duplex prior to use. More importantly, several studies have showed that sgRNA has better stability than crRNA:tracrRNA when duplexed with Cas9, thus leading to higher editing efficiency^1,2.

1. Hendel, et al., Chemically modified guide RNAs enhance CRISPR-Cas genome editing in human primary cells. Nat. Biotechnol., 33 (2015) 985-989.
2. Ryan et al., Improving CRISPR–Cas specificity with chemical modifications in single-guide RNAs. Nucleic Acids Research, 46 (2018) 2: 792–803.
What is the advantage of your sgRNAs compared to sgRNA synthesized from in vitro transcription?

Synthesis of 100 nt long sgRNAs was traditionally possible through in vitro-transcription (IVT) using phage RNA polymerase. These in vitro transcribed sgRNAs contain a 5’-triphosphate, which was thought to trigger immune response in many cell types. A recent study showed that sgRNAs with 5’-triphosphate modifications produced through in vitro-transcription can indeed induce innate immune responses and lead to cytotoxicity in human and murine cells. However, chemically synthesized sgRNAs without the 5’-triphosphate modifications demonstrated much better editing efficiency in cells, thus supporting that chemically synthesized sgRNAs are the most ideal reagent for CRISPR genome editing up till now³.

3. Kim et al. CRISPR RNAs trigger innate immune responses in human cells. Genome Res. 2018. 28: 367-373.
Why should I choose modified sgRNA versus unmodified versions?

Our modified sgRNA has 2’-O-methyl and phosphorothioate modifications at the first three 5’ and 3’ terminal RNA residues.2′ O-Methyl oligo modification is best characterized as a RNA analog which offers stability against hydrolysis and nucleases. The phosphorothioate (PS) modification renders the internucleotide linkage more resistant to nuclease degradation.

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CRISPR Synthetic sgRNA

Overview

Advantages