上海金畔生物科技有限公司代理New England Biolabs(NEB)酶试剂全线产品,欢迎访问官网了解更多产品信息和订购。 产品信息 EnGen® Spy Cas9 切刻酶是 Cas9 核酸酶的突变体,其在 RuvC 核酸酶结构域中引入了点突变(D10A),因而可以产生切刻,但不能切割 DNA(1,2)。与 EnGen Cas9 NLS 切刻酶(NEB #M0646)...
上海金畔生物科技有限公司代理New England Biolabs(NEB)酶试剂全线产品,欢迎访问官网了解更多产品信息和订购。 产品信息 EnGen®Spy dCas9 核酸酶是 Cas9 核酸酶的无活性突变体,保留了与 DNA 的结合活性(1,2)。该核酸酶包含 D10A 和 H840A 这两个点突变,可分别导致 RuvC 与 HNH 核酸酶结构域失活。N 端 ...
9.如图2所示,以pjg090为模板,在扩增中使用了含有用于golden gate克隆的接头(ojh307和ojh308)的引物。推荐的试剂如下:1μl pcr产物,50ng pjg112、1μl cutsmart buffer(neb),0.4μl t4连接酶缓冲液(neb),5u bsa i(neb),20u t4dna连接酶(neb),并将ddh2o补齐至10μl。将反应孵育20～25个循环(37℃2...
cas9/cas9d10amrna及sgrnas的体外转录操作各步骤均按相应试剂盒所述方法操作。简述如下:pst1374-cas9/cas9d10a-nls质粒经plasmidmidi试剂盒(qiagen,12143)提取,agei-hf(neb,r3552)消化,pcr纯化试剂盒(qiagen,28004)纯化后用mmessagemmachinet7ultrakit(ambion,lifetechnologies,am1345)体外转录获得cas9mrna。cas9mrna...
Cas9 H840A nickase was purchased from Applied Biological Materials Inc. (K136). Cas9 D10A nickase was purchased from NEB (M0650T). ATTO550 labeled tracrRNA was purchased from IDT. crRNA was transcribed in vitro using HiScribe™ T7 Quick High Yield RNA Synthesis Kit (NEB, E2050S). ...
BsaI (NEB, R0535S) AgeI (NEB, R0552S) DraI (TAKARA, D1037A) T4 DNA Ripid ligation Kit(NEB,M2200S) PMSG (Sansheng, China,50IU/ml in normal saline, Aliquot and store at -80℃) HCG (Sansheng, China,50IU/ml in normal sa...
D10A and H840A are mutations that are sufficient to abolish the nuclease activity of S. pyogenes SF370 Cas98,28. Surveyor. Targeted loci were amplified by PCR using Phusion Flash (NEB) or Heruclase II (Agilent) polymerases and primers listed in Table S2. For sg17 and sg21, two ...
D10A and H840A are mutations that are sufficient to abolish the nuclease activity of S. pyogenes SF370 Cas98,28. Surveyor. Targeted loci were amplified by PCR using Phusion Flash (NEB) or Heruclase II (Agilent) polymerases and primers listed in Table S2. For sg17 and sg21, two ...
Unlike dCas9, nCas9 (D10A) nicking of the target strand stimulates cellular repair mechanisms, which leads to increased editing frequencies by both cytosine and adenosine base editors in the absence of a DSB. 1Center for Genome Engineering, Institute for Basic Science, Daejeon, Republic of ...
(d)通过在指示的温度孵育5min后进行的活性测定,比较thermocas9和spcas9的活性温度范围。添加预加热的dna底物,并且将反应在相同的温度孵育持续1小时。 图18示出了在嗜热菌中的基于thermocas9的基因组工程。 (a)基本的pthermocas9_δ感兴趣基因(goi)构建体的示意图。引入thermocas9基因至pnw33n(史氏芽孢杆菌)或pemg...