DatasheetforSNU-475/Cas9-hygStableCellLine
Catalognumber:
SL522
Product:
SNU-475celllinestablyexpressingCRISPRCas9nuclease
Description:
ThisproductisacelllinestablyexpressingCRISPRCas9nuclease.Thiscellline
alsoexpressesthehygromycinresistancegene.Incombinationwithseparately
transfectedortransducedsingleguideRNAs(sgRNAs),thiscelllinewillsustain
double-strandDNAbreaks(DSBs)attargetedgenomesites.Thiscelllinecanbe
used in vitro for gene knockout, transgene knock-in, mutagenesis, transgene
integration,orothergenomeediting-relatedapplications.
Quantity:
1vialof2x106 cells;frozen
Shippingconditions:Dryice
Storage conditions: Liquid nitrogen vapor phase. Remove the item from the dry ice packaging andcheckallitemsfordamageandleakage.Placeimmediatelyintostorageat
orbelow-1400C,preferablyintotheliquidnitrogenvaporphase,untiluse.
Transgeneintegration:
Sourceofparentalline:
SNU-475
Organism: HumanTissue: Disease:
Celltype: Homosapiens,
Liver
Hepatocellularcarcinoma
Epithelial
Safetyinstructions: Toensuresafety,protectivegloves,clothing,andafacemaskshouldbeworn
whenhandlingfrozenvials.Someleakagemayoccurintothevialduring
storage.Theliquidnitrogenwillbeconvertedtogasuponthawing.Duetothe
natureofnitrogengas,pressuremaybuildwithinthevialandpossiblyresultin
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thevialexplodingorlosingitscap.Thismaycauseflyingdebris.
Thawingprocedure: Thevialofcellsshouldbethawedina37 0Cwaterbathwithgentleagitation.
For optimal performance, the vial should be thawed in under two minutes.
Ensurethatthecapofthevialdidnotloosenuponthawing,andre-tightenas
needed.Spraythevialwith70%EtOHandwipeoffbeforetransferringthevial
intocellculturehood.Usingaseptictechnique,addthecontentsofthevialto9
ml of complete growth medium (without selection), centrifuge for 5 minutes
at 250 x gtoremovethecryoprotectivemedium.Resuspendthecellpelletin
10 mL of complete growth medium, and place into a culture vessel of your
choice.Onlyaddselectiontothemediumafter24hoursinculture.
Cultureconditions:
CompleteGrowthMedium
ThebasemediumforthiscelllineisRPMI.Foroptimalgrowthandmaintenance
of selection, add the following components to the base medium: Fetal bovine
serumtoafinalconcentrationof10%.
Selection
Hygromycintoafinalconcentrationof150µg/mL
Culturetemperature:
370Cwith5%CO2
Subculture:
RinsethecellswithPBSwithoutcations,digestcellswith0.25%(w/v)TrypsinEDTA(0.53mM)solutionandsplitat1:2to1:4ratio.
Cryopreservation:
Freezeslowlyincompletegrowthmediumsupplementedwith5%(v/v)DMSO.
ProductQC:
>95%viabilitybeforefreezing.Allcellsweretestedandfoundtobefreeof
mycoplasma,bacterial,viruses,andothertoxins.
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Cas9ActivityTestingbyT7EndonucleaseI(T7E1)Assay
TE1:T7E1(T7Endonuclease1)
sgRNAtargetingEMX1genewastransducedinto
SNU-475/Cas9-hygStableCellLineby
transduction.EMX1genewascutbyCAS9
expressedinsidethecellsandrepairedthrough
NHEJwithmutation.A684bpEMX1gene
fragmentfromPCRwasthentestedbyT7
EndonucleaseI(T7E1)Assay.TheT7E1cleavage
willresultintwoadditionalbands:
one~315bpandtheother~369bp.
Citationofproduct: If use of this item results in a publication, please use this information:
CRISPRCas9SNU-475/Cas9-hygStablecellline(SL522,GeneCopoeia,Inc.,
Rockville,MD).
LimitedUseLicense
AlimiteduselicenseisgrantedtotheBuyeroftheProduct.TheProductshallbeusedbytheBuyerforinternal
researchpurposesonly.TheProductisexpresslynotdesigned,intended,orwarrantedforuseinhumansorfor
therapeuticordiagnosticuse.TheProductmustnotberesold,repackagedormodifiedforresale,orusedto
manufacturecommercialproductswithoutpriorwrittenconsentfromGeneCopoeia.ThisProductshouldbeused
inaccordancewithNIHguidelinesdevelopedforrecombinantDNAandgeneticresearch.Useofanypartofthe
Productconstitutesacceptanceoftheaboveterms.
Copyright©2016GeneCopoeia,Inc.
C9SCL-DS-102816-1
GeneCopoeia,Inc.
9620MedicalCenterDrive,#101
Rockville,MD 20850USA
Tel:301-762-0888;Fax:301-762-3888
Email:[email protected]
Web:www.genecopoeia.com
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