Product Name: Cas9 mRNA
Product Type: Chemical
CAS NO: 244240-24-2SGK inhibitors
Shipped in: dry ice
Storage temp.: −70°C
Application: Functional Genomics/Transgenic Applications
•Creation of gene knockouts in transgenic applications
•Creation of knock-in animals with promoters, fusion tags or reporters integrated into endogenous genes
Components:
1 vial containing 25ug of Cas9 mRNA.
Please note, product does not contain guideRNA sequence. This must be purchased separately through the Custom CRISPR product tab.
Features and Benefits:
RNA ready to use in transgenic applications. Must be used in conjunction with a purified guideRNA sequence to mediate a site specific double strand break in the DNA.
General description: Cas9 mRNA generated through in-vitro transcription which has been capped and polyA-tailed. This mRNA should be used in conjunction with purified guideRNA. The use of RNA vs. plasmid constructs avoids complications with promoter-embryo compatibility as well as the possibility of random integration of nuclease and gRNA-expressing plasmids into the host animal genome.
Legal InFormation: CRISPR Patents Pending
Other Notes:
Must be used in conjunction with the RNA Format of a guideRNA sequence in order to mediate a double strand break in the DNA.
Other Notes:
Typical microinjection concentrations used in the literature are in the ranges of 20-200 ng/ul for Cas9 mRNA and 10-50 ng/ul for gRNA.
Other Notes:
While Sigma CRISPR RNA was not developed for robust application to cell culture, we have seen some success in detecting CEL-I activity using RNA-only Formats for both CRISPR nucleases and paired nickases. RNA-only delivery Formats may be favorable for cell types which are sensitive to double stranded DNA such a dendritic cells or when promoter-cell incompatibilities exist.
Physical Form: Sigma Cas9 mRNA is supplied at a concentration of 500 ng/ul (50 ul Cas9 mRNA, capped and polyA-tailed)
Preparation Note: While Sigma implements the same high quality RNA synthesis procedures that have been successful in many mRNA-based ZFN transgenic projects, we highly recommend centrifuging your final, concentration-adjusted CRISPR RNA samples immediately prior to microinjection to ensure all particulate material has been removed which may result in clogging of microinjection needles.
Principle:
CRISPR/Cas systems are employed by bacteria and archaea as a defense against invading viruses and plasmids. Recently, the type II CRISPR/Cas system from the bacterium Streptococcus pyogenes has been engineered to function in eukaryotic systems using two molecular components: a single Cas9 protein and a non-coding guide RNA (gRNA). The Cas9 endonuclease can be programmed with a single gRNA, directing a DNA double-strand break (DSB) at a desired genomic location. Similar to DSBs induced by zinc finger nucleases (ZFNs), the cell then activates endogenous DNA repair processes, either non-homologous end joining (NHEJ) or homology-directed repair (HDR), to heal the targeted DSB.
RIDADR: NONH for all modes of transport
Storage Temp.: −70°C
UNSPSC
12352200
PubMed ID:http://jpet.aspetjournals.org/content/97/3/292