Code	제품명	대체품
631438	Guide-it™ sgRNA In Vitro Transcription Kit	632635
631439	Guide-it™ Complete sgRNA Screening System	632636
631440	Guide-it™ sgRNA Screening Kit	632639

CRISPR/Cas9을 이용하여 genome modification 하기 위한 single guide RNA(sgRNA)의 간편한 합성 및 스크리닝이 가능한 제품군
In vitro transcription을 통해 sgRNA 4 μg 이상 합성 가능
Cell 도입하기 전에 간편하게 sgRNA효율 확인 가능
고품질의 Cas9 nuclease 사용

제품설명

Guide-it Complete sgRNA Screening System (Code 631439)은 sgRNA합성과 효율을 확인할 수 있는 제품이다. 본 제품은 sgRNA 합성 kit와 효율 확인 kit로 구성된다.
Guide-it sgRNA In Vitro Transcription Kit (Code 631438)는 CRISPR/Cas9 genome editing에 사용하는 sgRNA을 in vitro transcription 방법으로 합성하고, 간편하게 높은 비율에 sgRNA를 만들 수 있다.
Guide-it sgRNA Screening Kit (Code 631440)는 합성한 sgRNA의 효율을 확인하는 제품이다. 목적 세포로 도입 전에 sgRNA의 효과를 확인할 수 있어 시간과 노력을 절약할 수 있다.

Figure 1. Using Guide-it technology to perform genome editing. A sgRNA consists of a crRNA sequence that binds to a specific DNA target, and a tracRNA sequence that binds to Cas9 protein. When an sgRNA engineered to contain a target sequence of interest binds to a recombinant form of Cas9 nuclease that has double-stranded DNA nicking activity, the resulting complex will produce target-specific double-stranded cleavage. Cellular repair, which is error-prone, will take place at the cleavage site, and may result in a mutation that can "knock out" a gene.

Figure 2. The Guide-it Complete sgRNA Screening System protocol.

Figure3. Overview of Guide-it Complete sgRNA Screening System

Figure 4. Analysis of CRISPR Cas9 cleavage products. A guide RNA (sgRNA) sequence was synthesized and tested against its target in human chromosome 10. A 2 kb fragment of chromosome 10 was PCR-amplified, and an sgRNA designed to cleave the amplified sequence was in vitro-transcribed using the Guide-it Complete sgRNA Screening System (Code 631439). The chromosome 10 fragment, the sgRNA, and recombinant Cas9 enzyme (also included in the kit) were combined in an in vitro cleavage reaction according to the protocol. Agarose gel analysis indicated that 10 ng (Lane 3) and 20 ng (Lane 4) samples of sgRNA were each sufficient for 100% cleavage of the target fragment. Lane 2 shows the untreated/uncleaved control.

구성품 (자세한 내용은 CoA를 참조하세요)

Guide-it Complete sgRNA Screening System (Code 631439)
1 each	Guide-it sgRNA In Vitro Transcription Components (Code 631441; 10 rxns; not sold separately)
	50 μl	Guide-it Scaffold Template (20 ng/μl)
	70 μl	Guide-it In Vitro Transcription Buffer
	30 μl	Guide-it T7 Polymerase Mix (33 U/μl)
	500 μl	RNase Free Water
1 each	Guide-it Recombinant Cas9 Nuclease (Code 631442; 30 rxns; not sold separately)
	30 μl	Guide-it Recombinant Cas9 Nuclease (500 ng/μl)
	30 μl	10X Cas9 Reaction Buffer
	30 μl	10X BSA
	500 μl	RNase Free Water
	10 μl	2 kb Control Fragment (40 ng/μl)
	6 μl	Control sgRNA (20 ng/μl)
1 each	NucleoSpin Gel and PCR Clean-Up (Sample) (Code 740609.10; 10 preps; not sold separately)
1 each	High Yield PCR EcoDry Premix (Code 639278; 24 rxns)
*Guide-it sgRNA In Vitro* Transcription Kit (Code 631438)**
1 each	Guide-it sgRNA In Vitro Transcription Components (Code 631441; 10 rxns; not sold separately
1 each	NucleoSpin Gel and PCR Clean-Up (Sample) (Code 740609.10; 10 preps; not sold separately)
1 each	High Yield PCR EcoDry Premix (Code 639278; 24 rxns)
Guide-it sgRNA Screening Kit (Code 631440)
1 each	Guide-it Recombinant Cas9 Nuclease (Code 631442; 30 rxns; not sold separately)
1 each	NucleoSpin Gel and PCR Clean-Up (Sample) (Code 740609.10; 10 preps; not sold separately)
1 each	High Yield PCR EcoDry Premix (Sample) (Code 639279; 8 rxns)

보존

Control sgRNA, Guide-it Recombinant Cas9 Nuclease : -70 ℃
NucleoSpin Gel and PCR Clean-up : 실온
High Yield PCR EcoDry Premix : 실온(20~22 ℃ : 건조기 내에서 보존)
기타 : -20℃