Overview

Double-stranded DNA (dsDNA) has been used for CRISPR homology directed repair (HDR) template for creating gene knock-in with high editing efficiency. GenWand™ dsDNA is developed with covalently closed ends to mitigate no-homologous end joining risk and increase knock-in accuracy. GenScript now offers up to g level closed-end dsDNA ideal for screening, process optimization and scale up.

Why closed-end dsDNA as CRISPR Gene Knock-In HDR Templates?

  • Lower toxicity and higher KI efficiency compare to PCR product
  • Closed-end to mitigate NHEJ events
  • No introduction of unnecessary plasmid sequences
  • Ideal for people need to KI long sequence
  • Ideal for large scale screening and scale up

Why GenWand™ dsDNA Service?

  • Made via plasmids based production process
  • 2-10 kb
  • µg to g level production
  • Comprehensive QC to ensure minimum impurities
  • Built in endotoxin removal process and guaranteed < 10 EU/mg Endotoxin
  • Quality grades designed for different research stage (RUO, Preclinical, Basic GMP Grade)

Services

dsDNA

Starting at 50µg/item , Deliver in as fast as 3 weeks .

Length (bp) Research Grade Preclinical Grade
2000~5000 Starting from $1,800/item Starting from $2,050/item
5001~10000 Starting from  $2,200/item Starting from $2,500/item

Test Specifications Detection Method Release Criteria Research Grade Preclinical Grade
Purity Agarose gel electrophoresis Single band
Sequence accuracy Sanger sequencing 100% sequence alignment
Optical density Spectrophotometer at 260 nm/230 nm ≥ 2.0
Spectrophotometer at 260 nm/280 nm 1.8~2.0
Bioburden Incubation in TSA plate No colony formation -
Endotoxin Qualitative TAL assay < 10 EU/mg -
Protein Residue Micro BCA Protein Assay Kit ≤50µg/mg -
Quantitative purity Agilent 2100 Bioanalyzer ≥90% - Add-on
pH pH meter with pH probe Depending on buffer pH - Add-on
Conductivity pH meter with conductivity probe Depending on buffer conductivity - Add-on

Applications

CRISPR based gene insertion, replacement, or correction

Mechanism of CRISPR HDR based gene editing

CRISPR/Cas9 technology is commonly used to create precise double stranded breaks (DSBs) at target DNA sites. The guide RNA (gRNA) recognizes the protospacer adjacent motif (PAM) sequence on the target DNA after forming complex with Cas9, then Cas9 exerts its endonuclease function to cause DSBs. This triggers two mechanisms for repair: one is non-homologous end-joining (NHEJ), which introduces mutations in the DSB site. The other mechanism is homology directed repair (HDR) which enables the donor DNA to be inserted at the break site and create gene knock-ins.

Resources

Get in Touch
with GenScript Custom
CRISPR Experts

Get in Touch with GenScript Custom CRISPR Experts
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