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Gene transfer (virus)
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Adeno-Associated Virus (AAV)
> CRISPR/SaCas9 Vector System (AAV2)
SaCas9¸¦ ÀÌ¿ëÇÑ AAV mediated CRISPR Genome Editing (Single vector)
CRISPR/SaCas9 Vector System (AAV2)
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Clontech
632619
AAVpro¢ç CRISPR/SaCas9 Helper Free System (AAV2)
1 Kit
(Package)
1,674,000¿ø
,
Clontech
632618
AAVpro¢ç CRISPR/SaCas9 Vector System
1 Kit
(Package)
789,000¿ø
,
Product index
Adeno-Associated Virus (AA..
Overview - All about A..
Packaging (293T Cell L..
AAV1, 2, 5, 6 Vector S..
CRE Recombinase System
LacZ System
pAAV-ZsGreen1 Vector
CRISPR/Cas9 Vector Sys..
CRISPR/SaCas9 Vector S..
Tet-Inducible System (..
Cell & Sup. Purificati..
Purification Kit Maxi/..
Purification Kit (AAV2..
AAVpro¢ç Concentrator
AAVpro¢ç Titration Kit..
Extraction Solution
AAV Freeze-thaw extrac..
Lentivirus
[°³¿ä] Packaging Singl..
[FAQ] Lentiviral Gene ..
CMV promoter º¤ÅÍ ½Ã½º..
EF1 ¥á promoter º¤ÅÍ ..
Lenti-X¢â SARS-CoV-2 P..
3¼¼´ë Lentivirus packa..
Çü±¤´Ü¹éÁú ¸®Æ÷ÅÍ º¤ÅÍ
Tet-on inducible Expre..
Tet One Inducible Expr..
Lenti-X¢â CRISPR/Cas9 ..
(Legacy) Tet-Inducible..
Reporter assay ½Ã½ºÅÛ ..
Packaging single shots
Packaging Cell Lines
Lenti-X¢â qRT-PCR Titr..
Lenti-X¢â p24 Rapid Ti..
GoStix¢â Plus
Lentivirus ³óÃà ½Ã¾à
Lentivirus Á¤Á¦
Ecotropic Receptor Boo..
RetroNectin¢ç (Recombi..
Transduction ½Ã¾à
Provirus Quantitation ..
Integration Site Analy..
Lenti-X¢â Transduction..
TransIT¢ç Lentivirus S..
Retrovirus
Retro-X ¼±Åà °¡À̵å
RetroNectin¢çÀÇ ´Ù¾çÇÑ..
Packaging Cell £¦ ½Ã½º..
Retrovirus vector £¦ ..
Çü±¤´Ü¹éÁú Retrovirus ..
MSCV Retrovirus ½Ã½ºÅÛ
Retro-X Q vector
pDON-5 DNA
pDON-AI-2 DNA
pMEI-5 DNA
Retrovirus Packaging K..
Tet À¯µµ¹ßÇö ½Ã½ºÅÛ (T..
Tet One Inducible Syst..
qRT-PCR titer ÃøÁ¤
Titer Set (for Real Ti..
Retrovirus ³óÃà
Receptor Boosters (Eco..
Provirus Copy ¼ö ÃøÁ¤
Integration Site Analy..
Adenovirus
Adeno-X¢â workflow
Adenoviral Expression ..
Packaging Cell Lines
Adenovirus Á¤Á¦ (Maxi ..
Adenovirus Á¤Á¦ (Mega ..
qPCR titer ÃøÁ¤ Kit
Hexon ±â¹Ý titer ÃøÁ¤
Adeno-X GoStix
CAR Receptor Booster
Mammalian Cell ¿ë Vector
[¼±Åð¡À̵å] ´Ù¾çÇÑ Á¾..
¾ç¹æÇ⼺ ¹ßÇö º¤ÅÍ (Bi..
EF1¥á Promotor Vector ..
epitope tag (HA or c-M..
in vivo Vector, pLIVE ..
Magnetic Myc IP Kit
pBApo-CMVvector ½Ã¸®Áî
pBApo-EF1¥á Vector ½Ã..
pIRES Bicistronic Expr..
¡Ø SpCas9 (from
S.pyogenes
) ¸¦ ÀÌ¿ëÇÑ AAV meditated CRISPR/Cas9 Genome EditingÀº
AAVpro¢ç CRISPR/Cas9 Helper Free System (AAV2) (Code 632608)
(Two vector system)À» È®ÀÎÇϼ¼¿ä.
Staphylococcus aureus
À¯·¡ÀÇ SaCas9¸¦ ÀÌ¿ëÇÑ One vector CRISPR/SaCas9 System
Adeno-associated virus (AAV)¸¦ ÀÌ¿ëÇÏ¿© SaCas9°ú sgRNA¸¦ ¼¼Æ÷ ³» µµÀÔ
TransfectionÀÌ ¾î·Á¿î Mammlian cell (ºÐ¿, ºñºÐ¿¼¼Æ÷)¿¡¼ Genome Editing
AAV¿¡ ÀÇÇÑ Cas9 µµÀÔÀº genome integrationÀ» ¾ïÁ¦ÇÏ°í Cas9ÀÇ ¹ßÇöÀ» Áö¼Ó½Ãų »Ó¸¸ ¾Æ´Ï¶ó, off-target effect¸¦ ¾ïÁ¦
Á¦Ç°¼³¸í
AAVpro¢ç CRISPR/SaCas9 Helper Free SystemÀº single guide RNA (sgRNA)¿Í SaCas9 (from
Staphylococcus aureus
) À¯ÀüÀÚ¸¦ ¹ßÇöÇÏ´Â Adeno-associated virus 2 (AAV2) particles¸¦ Helper virus ¾øÀÌ Á¦ÀÛ °¡´ÉÇÑ Á¦Ç°ÀÌ´Ù. º» Á¦Ç°Àº
S.aureus
À¯·¡ÀÇ SaCas9°ú sgRNA¸¦ ÇϳªÀÇ º¤ÅÍ¿¡¼ ¹ßÇöÇÒ ¼ö ÀÖ´Â single vector systemÀÌ´Ù (±×¸² 1).
S.aureus
À¯·¡ÀÇ SaCas9 nucleaseÀÇ À¯ÀüÀÚ Å©±â´Â 3.3 kb·Î½á, ±âÁ¸¿¡ »ç¿ëµÇ¾î¿Ô´ø
Streptococcus pyogenes
À¯·¡ÀÇ SpCas9 nuclease (4.1 kb) (º¸Åë Cas9À̶ó°í ¸í¸íÇÔ) º¸´Ù ¾à 1 kb Á¤µµ ÀÛ´Ù. ±×·¸±â ¶§¹®¿¡ ÇϳªÀÇ AAV vector¿¡ sgRNA ¹ßÇö À¯ÀüÀÚ¿Í SpCas9 ¹ßÇö À¯ÀüÀÚ¸¦ ÇѲ¨¹ø¿¡ »ðÀÔÀÌ °¡´ÉÇÏ°Ô µÇ¾úÀ¸¸ç, ÀÌ´Â
in vitro
¹×
in vivo
¿¡¼ ´Ù¾çÇÑ ¼¼Æ÷¿¡¼ È¿À²ÀûÀ¸·Î Genome editingÀÌ °¡´ÉÇÏ´Ù. ¹Ý¸é, SaCas9ÀÇ °æ¿ì SpCas9°ú PAM ¼¿ (Proto-Spacer Adjacent Motif) ¹× target sequenceÀÇ ±æÀÌ (20 - 22 nt)°¡ ´Ù¸£¹Ç·Î sgRNA µðÀÚÀÎ ´Ü°è¸¦ °í·ÁÇÒ ÇÊ¿ä°¡ ÀÖ´Ù (±×¸² 2).
º» Á¦Ç°¿¡ Æ÷ÇԵǾî ÀÖ´Â SaCas9/sgRNA ¹ßÇö AAV vector, AAV packaging¿¡ ÇÊ¿äÇÑ pRC2-mi342 vector ¹× pHelper vector¸¦ HEK293T ¼¼Æ÷ÁÖ¿¡ µµÀÔÀ» Çϸé, Helper virus ¾øÀÌ °í¼øµµ ¹× ³ôÀº TiterÀÇ AAV particleÀ» ¾òÀ» ¼ö ÀÖ´Ù. º» Á¦Ç°¿¡´Â AAV production ¹× CRISPR Genome editingÀ» À§ÇÑ ¸ðµç Á¦Ç° (Ligation mix, Stellar¢â Competent cells, AAV Extraction buffer µî)ÀÌ Æ÷ÇÔ µÇ¾îÀÖ´Ù.
±×¸² 1. pAAV-Guide-it-1 vectorÀÇ SaCas9°ú sgRNA ¹ßÇö
pAAV-Guide-it-1 vector¸¦ ÆÐŰ¡ÇÏ°í ÀÖ´Â AAV particleÀ» Target cell line¿¡ transductionÇÔÀ¸·Î½á, SaCas9 ´Ü¹éÁú°ú sgRNA À¯ÀüÀÚ°¡ µ¿½Ã¿¡ ÇÔ²² ¹ßÇöµÈ´Ù.
±×¸² 2. SaCas9ÀÇ PAM sequence
SaCas9ÀÇ PAM ¼¿Àº NNGRR (N=A, T, G, C / R=A/T)À̸ç, PAMÀ¸·ÎºÎÅÍ 3 - 4 bpÀÇ ¿°±â¸¦ Àý´ÜÇÑ´Ù. ¶ÇÇÑ, sgRNAÀÇ Target sequence´Â 20 - 22nt¸¦ ±ÇÀåÇÑ´Ù. SaCas9ÀÇ sgRNA µðÀÚÀÎÀº
http://benchling.com
¿¡¼ °¡´ÉÇÏ´Ù.
±×¸² 3. AAV¸¦ ÀÌ¿ëÇÑ SaCas9°ú SpCas9ÀÇ Genome editing È¿À² ºñ±³
CYP2
Exon1 À¯ÀüÀÚ Áß, 2°³ÀÇ ´Ù¸¥ ¹è¿À» editing Ç¥ÀûÀ¸·Î ÇÏ¿© Two vector systemÀÎ
AAVpro¢ç CRISPR/Cas9 Helper Free System (AAV2) (Code 632608)
°ú Sinlge vector systemÀÎ
AAVpro¢ç CRISPR/SaCas9 Helper Free System (AAV2) (Code 632619)¸¦ »ç¿ëÇÏ¿© AAV mediated CRISRP/Cas9 editingÀ» ¼öÇàÇÏ¿´´Ù. 12-well plate¿¡ 1 x 10
5
°³ÀÇ ¼¼Æ÷¸¦ seeding, over night ¹è¾ç ÈÄ, SpCas9 or SaCas9 virus vector¸¦ 1 x 10
5
MOI·Î °¨¿°½ÃÄ×´Ù. 72½Ã°£ ÈÄ, ¼¼Æ÷¸¦ ȸ¼öÇÏ°í Genome DNA º¯À̵µÀÔ È¿À²À»
Guide-it¢â Mutation Detection Kit (Code 631443)
À» »ç¿ëÇÏ¿© ºÐ¼® ¹× ºñ±³ÇÏ¿´´Ù.
Applications
AAV-based delivery of a user-defined sgRNA and Cas9 for mammalian genome editing using CRISPR/Cas9 technology
±¸¼ºÇ° (ÀÚ¼¼ÇÑ ³»¿ëÀº CoA¸¦ ÂüÁ¶Çϼ¼¿ä)
AAVpro¢ç CRISPR/SaCas9 Helper Free System (AAV2) (Cat. No. 632619)
AAVpro¢ç CRISPR/SaCas9 Vector Set
Guide-it¢â Ligation Components
Stellar¢â Competent Cells
pRC2-mi342 Vector
pHelper Vector
AAVpro¢ç Extraction Solution
AAVpro¢ç CRISPR/SaCas9 Vector System (Cat. No. 632618)
AAVpro¢ç CRISPR/SaCas9 Vector Set
Guide-it¢â Ligation Components
Stellar¢â Competent Cells
º¸Á¸
Stellar¢â Competent Cells: -80 ¡É
±âŸ: -20 ¡É
(AAVpro¢ç Extraction SolutionÀº À¶ÇØ ÈÄ, ½Ç¿Â º¸°ü)
Keyword :
Adeno-Associated Virus (AAV)
Overview - All about AAV
Adeno-associated virus Á¦ÀÛ¿¡ ÃÖÀûÀÎ packagi..
Packaging (293T Cell Line)
Helper Free ½Ã½ºÅÛ¿¡ ÀÇÇÑ ´Ù¾çÇÑ serotypeÀÇ ..
AAV1, 2, 5, 6 Vector System
Cre-Recombinase À¯ÀüÀÚ¸¦ žÀçÇÑ AAV1, AAV2, ..
CRE Recombinase System
¥â-galactosidase À¯ÀüÀÚ¸¦ žÀçÇÑ AAV1, AAV2,..
LacZ System
Çü±¤´Ü¹éÁúÀÌ Æ÷ÇÔµÈ AAV vector
pAAV-ZsGreen1 Vector
Adeno-associated virus delivery-CRISRP/Cas9 ..
CRISPR/Cas9 Vector System
SaCas9¸¦ ÀÌ¿ëÇÑ AAV mediated CRISPR Genome E..
CRISPR/SaCas9 Vector System (AAV2)
AAV2 Delivery Tet À¯µµ¹ßÇö ½Ã½ºÅÛ
Tet-Inducible System (AAV2)
Cell°ú ¹è¾ç ¹èÁö¿¡¼ AAV ÃßÃâºÎÅÍ Á¤Á¦±îÁö
Cell & Sup. Purification Kit
´Ù¾çÇÑ serotypeÀÇ AAV Á¤Á¦¿¡ ÃÖÀû
Purification Kit Maxi/Midi (All Serotypes)
AAV Serotype 2 Á¤Á¦ kit
Purification Kit (AAV2 Àü¿ë)
AAV¸¦ È¿À²ÀûÀ¸·Î ³óÃà
AAVpro¢ç Concentrator
ºü¸£°í Á¤È®ÇÑ AAV ¿ª°¡ÃøÁ¤ (qPCR)
AAVpro¢ç Titration Kit (for Real Time PCR)
°£ÆíÇÑ AAV ÃßÃâ½Ã¾à
Extraction Solution
±âÁ¸ÀÇ AAV ÃßÃâ ¹æ¹ýÀ¸·Î Á¤Á¦±îÁö 4½Ã°£ ³» ..
AAV Freeze-thaw extraction solution