基因組編輯工具-CRISPR/Cas9
繼ZFN（Zinc Finger Nucleases）技術后，Merck在2013年推出新一代基因組編輯工具--CRISPR/Cas9，讓研究人員以更快、更經濟的方式實現基因組特定位點的編輯。

基因組編輯工具-CRISPR/Cas9

繼ZFN（Zinc Finger Nucleases）技術后，Merck在2013年推出新一代基因組編輯工具--CRISPR/Cas9，讓研究人員以更快、更經濟的方式實現基因組特定位點的編輯。憑借過去10年在基因組編輯領域的豐富經驗積累以及專業的生物信息學平臺，Merck已經成功設計出覆蓋人類，小鼠和大鼠三個物種的所有基因的CRISPR/Cas9載體，并可以提供在線定制服務，以及完整的CRISPR實驗workflow解決方案。此外，默克與Sanger Institute合作開發了人、小鼠全基因組CRISPR 文庫，以幫助科學家實現基因功能的快速篩選、規模化的模型建立以及藥物作用篩選等。

• 高效：優化的載體設計，大限度提高轉染效率，簡化篩選工作
• 特異：特殊的gRNA設計和雙切口酶系統，大限度提高特異性
• 全面：產品齊全，可提供質粒、RNA、慢病毒載體、RNP等形式，涵蓋人、大鼠、小鼠、植物等多個物種，更有Sanger Arrayed和Broad Pools全基因組文庫以及重要通路的亞文庫
• 掌控：強大的慢病毒全基因組文庫可輕松進行高通量篩選，全面掌控人或小鼠的基因組

 

CRISPR實驗--載體構建和表達

• CRISPR/Cas gRNA設計和選擇 • crRNA定制合成 • 克隆和表達 • 抗生素 • GenElute™質粒DNA快速提取kit • PCR擴增和核酸電泳 • SP6/T7 體外轉錄試劑盒 • GenElute™總RNA提取試劑盒 • 慢病毒包裝系統 • 培養皿/離心管/槍頭等基礎耗材

Molecular Biology Cloning & Expression

The aim of molecular cloning is to insert the gene-of-interest (GOI) into a plasmid vector. This vector is then inserted into a cell that will express the protein encoded by the GOI. Once protein is expressed, the protein function can be studied as it affects the cell signaling, morphology or other aspects. Alternatively, the protein can be expressed in large quantities that can then be studied directly with other techniques. We have a wide variety of products for use across the cloning & expression workflow.

expression workflow.

 

Cloning Products

• SnapFast™ Vectors • Cloning Enzymes • Restriction Enzymes • Biomatrica™ Nucleic Acid Stabilization • Custom DNA Oligos

Restriction Enzyme Buffer Reference

Sigma^® Life Science’s restriction enzyme collection has been optimized for digestion using five unique buffers. Reference below the enzyme activity in each buffer (table 1), and the preferred buffer for each enzyme (table 2).

 

Table 1
Activity (%) of each Sigma restriction enzyme in each Sigma buffer. The preferred buffer for each enzyme is highlighted in bold.
 

	Buffer
Restriction Enzyme	SA (B7531)	SB (B8781)	SL (B3782)	SM (B3158)	SH (B3657)
Acc I (R6142)	100	0-10	10-25	1-10	0-10
Alu I (R6885)	100	50-75	25-50	25-50	0-10
Apa I (R4258)	100	10-25	50-75	50-75	0-10
Bam HI (R0260)	100	100	75-100	100	25-50
Bcl I (R8631)	100	100	25-50	100	100
Bgl II (R6377)	100	100	25-50	100	100
Bln I (R3131)	25-50	50-75	0-10	25-50	100
Bsm I (R3635)	0-10	50-75	0-10	25-50	100
Cfo I (R1761)	75-100	50-75	100	50-75	25-50
Cla I (R7763)	100	100	75-100	100	100
Dde I (R4256)	50-75	75-100	25-50	25-50	100
Dpn I (R8381)	100	75-100	50-75	75-100	75-100
Dra I (R4381)	100	75-100	100	100	50-75
Ecl XI (R3884)	25-50	100	25-50	25-50	50-75
Eco RI (R6265)	100	100	25-50	50-75	100
Eco RV (R2756)	25-50	100	0-10	25-50	50-75
Hae III (R5628)	50-75	50-75	75-100	100	25-50
Hind III (R1137)	50-75	100	25-50	100	50-75
Hpa I (R8507)	100	25-50	25-50	50-75	25-50
Kpn I (R1258)	75-100	10-25	100	25-50	0-10
Nco I (R8761)	50-75	50-75	50-75	50-75	100
Nde I (R5509)	25-50	100	10-25	50-75	100
Nhe I (R5634)	100	25-50	100	100	10-25
Not I (R8506)	10-25	50-75	0-10	25-50	100
Nsi I (R5884)	50-75	100	10-25	50-75	100
Pst I (R7023)	25-50	25-50	10-25	25-50	100
Pvu I (R1508)	50-75	75-100	25-50	50-75	100
Pvu II (R2631)	25-50	25-50	25-50	100	25-50
Rsa I (R4756)	100	50-75	100	50-75	0-10
Sac I (R5268)	100	0-10	100	50-75	0-10
Sal I (R0754)	0-10	25-50	0-10	10-25	100
Sau 3A (R0762)	100	25-50	25-50	75-100	0-10
Sca I (R5007)	0-10	100	0-10	75-100	100
Sfi I (R8256)	25-50	25-50	75-100	100	25-50
Sma I (R4503)	100	0-10	0-10	0-10	0-10
Spe I (R5257)	75-100	75-100	75-100	100	100
Sph I (R7135)	50-75	75-100	25-50	100	75-100
Ssp I (R6759)	75-100	75-100	10-25	75-100	100
Taq I (R9507)	50-75	100	25-50	50-75	50-75
Xba I (R7260)	100	75-100	75-100	75-100	100
Xho I (R6759)	25-50	75-100	10-25	25-50	100

Table 2
The complete formulation of each Sigma Restriction Enzyme Buffer. The formulation of the storage buffer for each Sigma restriction enzyme is given on the catalog page (see links in table 1).
 

	Final concentration (mM)
	SA (B7531)	SB (B8781)	SL (B3782)	SM (B3158)	SH (B3657)
Tris-acetate	33
Tris-HCl		10	10	10	50
Mg-acetate	10
MgCl2		5	10	10	10
K-acetate	66
NaCl		100		50	100
Dithioerythritol (DTE)			1	1	1
Dithiothreitol (DTT)	0.5
2-Mercaptoethanol (BME)		1
pH at 37°C	7.9	8.0	7.5	7.5	7.5

Categories

• Buffers
• Digestions
• Enzyme activity
• PAGE
• Solid phase extractions