GlyScope系列产品

适用于糖链研究

◆背景

　　众所周知，糖链是通过结合蛋白（糖蛋白）和脂质（糖脂）在细胞中发挥功能。糖蛋白和糖脂已被证明在体内扮演多种多样的角色。

◆特点和优点

●　适用于实验室通用仪器

●　快速分析系统

●　可用于高效液相色谱法分析糖类 

TRPV4拮抗剂

【New】HC-067047

　　HC-067047是TRPV4（Transient Receptor  Potential Vanilloid 4）的强力拮抗剂。实验发现，TRPV4在低渗透压中是活性化的渗透压感受性和温度感受性受体。TRPV4还与皮肤的屏障功能有关。

　　HC-06704对TRPV1，V2，V3的抑制活性比对TRPV4的至少要高出100倍。对hEGF和TRPM8的抑制活性约为对TRPV4活性的十分之一。

◆ 产品简介

◆ 相关产品：

TRPV4激动剂

TRPV4拮抗剂

TRPV4抗体

   Enzo Life Sciences提供一系列独一无二的化合物库，包括各类抑制剂、激活剂和诱导剂等各种小分子。随货附送完整的证明书，化合物活性描述、包装规格、化合物浓度含量、物理性状和结构式。方便客户查阅。

◆优点、特色

u  广泛的产品线：

　  超过3000种小分子化合物，包括：天然化合物、酶抑制剂激活剂、手提配体、药物、脂肪酸等

　  独特的化合物库，包括：FDA认证化合物库、天然化合物库、化学基因组学和信号通路相关化学物库等。

u  新颖的化合物：

　  化合物库中包含各种小分子，包括Enzo公司最新研发的化合物。

u  简单、低成本：

　  化合物全部溶在相应溶剂中，无需额外溶解步骤，即可进行筛选。

Screen-Well^® 毒性化合物库

　Screen-Well^® 化合物库可用于心脏毒性、肝脏毒性和造血毒性研究，这些化合物毒性明确、对不同器官有相关毒性证明资料。该库中含有许多结构、机制不同的化合物，且有无毒化合物作为对照。该化合物库在预测毒理学筛选（包括高通量筛选）中非常有用。

◆优点、特色

u  大量毒性明确且毒性不同的化合物

u  包含无毒对照品

u  96孔板包装，溶于DMSO中，即可进行筛选

Natural Products

Natural product library

　天然产物化学多样性丰富，特别适用于筛选具有药理活性的小分子化合物。通常情况下，很多新药都是来源与天然产物。该化合物库中包括500多种已知结构的高纯度天然产物。化合物都溶于DMSO中。根据客户需要，部分化合物可提供大包装。

化合物包括：

丙烯酸自动干燥箱C型

通过风扇使空气循环对流，常温下柜内空气湿度保持在30～40％RH的低湿度。全自动无需更换硅胶。带锁柜门安全性高。

◆原理

◆优点・特色

◆案例・应用

干燥箱广泛应用于生物化学，化工制药、医疗卫生、农业科研、环境保护等研究应用领域，作粉末干燥、烘培以及各类玻璃容器的消毒和灭菌用。特别适合于对干燥热敏性、易分解、易氧化物质和复杂成分物品进行快速高效的干燥处理。

Phos-tag™ 琼脂糖吸管

Phos-tag™ Tip

　　可特异性捕捉磷酸基团的功能性分子“Phos-tag ™”的磷酸化肽纯化用枪头。

　　枪头里含有Phos-tag ™ 琼脂糖，是即开即用的前处理工具，适用于生理状态下低分子量磷酸化分子（核酸和多肽等）的分离和富集。

◆原理

把Phos-tagTM Tip 装在注射器（购买产品附送）上使用。

Phos-tag ™ Tip 的结构

◆优点、特色

●　操作时间少于30分钟。        ●　高回收率可重复利用。 ●　无需昂贵仪器。 ●　缓冲液为生理pH 条件下。

◆案例、应用

分离磷酸化肽使用例

Phos-tag™ 系列

磷酸化蛋白新方法！

　　Phos-tag™是一种能与磷酸离子特异性结合的功能性分子。它可用于磷酸化蛋白的分离（Phos-tag™ Acrylamide）、Western Blot检测（Phos-tag™ Biotin）、蛋白纯化 （Phos-tag™Agarose）及质谱分析MALDI-TOF/MS （Phos-tag™ Mass Analytical Kit）。

◆Phos-tag™ 的基本结构

◆原理：

◆特点：

●   与-2价磷酸根离子的亲和性和选择性高于其它阴离子

●   在pH 5-8的生理环境下生成稳定的复合物

◆相关应用：

◆相关产品：

phos-tag™由日本广岛大学研究生院医齿药学综合研究科医药分子功能科学研究室开发。

更多产品信息，请点击：http://phos-tag.jp

Phos-tag 第5版说明书

Phos-tag系列 ver 5

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·  Characterization of a herpes simplex virus 1 (HSV-1) chimera in which the Us3 protein kinase gene is replaced with the HSV-2 Us3 gene[J]. Journal of virology, 2016, 90(1): 457-473，Shindo K, Kato A, Koyanagi N, et al.

·  Generation of phospho‐ubiquitin variants by orthogonal translation reveals codon skipping[J]. FEBS letters, 2016, 590(10): 1530-1542，George S, Aguirre J D, Spratt D E, et al.

·  Evolution of KaiC-Dependent Timekeepers: A Proto-circadian Timing Mechanism Confers Adaptive Fitness in the Purple Bacterium Rhodopseudomonas palustris[J]. PLoS Genet, 2016, 12(3): e1005922，Ma P, Mori T, Zhao C, et al.

·  Phosphorylation of Bni4 by MAP kinases contributes to septum assembly during yeast cytokinesis[J]. FEMS Yeast Research, 2016, 16(6): fow060，Pérez J, Arcones I, Gómez A, et al.

·  Alteration of Antiviral Signalling by Single Nucleotide Polymorphisms (SNPs) of Mitochondrial Antiviral Signalling Protein (MAVS)[J]. PloS one, 2016, 11(3): e0151173，Xing F, Matsumiya T, Hayakari R, et al.

·  Arm-in-arm response regulator dimers promote intermolecular signal transduction[J]. Journal of bacteriology, 2016, 198(8): 1218-1229，Baker A W, Satyshur K A, Morales N M, et al.

·  The lsh/ddm1 homolog mus-30 is required for genome stability, but not for dna methylation in neurospora crassa[J]. PLoS Genet, 2016, 12(1): e1005790，Basenko E Y, Kamei M, Ji L, et al.

·  Fine tuning chloroplast movements through physical interactions between phototropins[J]. Journal of Experimental Botany, 2016: erw265，Sztatelman O, Łabuz J, Hermanowicz P, et al.

·  Characterization of the Neospora caninum NcROP40 and NcROP2Fam-1 rhoptry proteins during the tachyzoite lytic cycle[J]. Parasitology, 2016, 143(01): 97-113，Pastor-Fernandez I, Regidor-Cerrillo J, Jimenez-Ruiz E, et al.

·  Transcriptional Profile during Deoxycholate-Induced Sporulation in a Clostridium perfringens Isolate Causing Foodborne Illness[J]. Applied and environmental microbiology, 2016, 82(10): 2929-2942，Yasugi M, Okuzaki D, Kuwana R, et al.

·  Timely Closure of the Prospore Membrane Requires SPS1 and SPO77 in Saccharomyces cerevisiae[J]. Genetics, 2016: genetics. 115.183939，Paulissen S M, Slubowski C J, Roesner J M, et al.

·  DDK dependent regulation of TOP2A at centromeres revealed by a chemical genetics approach[J]. Nucleic Acids Research, 2016: gkw626，Wu K Z L, Wang G N, Fitzgerald J, et al.

·  OVATE Family Protein 8 Positively Mediates Brassinosteroid Signaling through Interacting with the GSK3-like Kinase in Rice[J]. PLoS Genet, 2016, 12(6): e1006118，Yang C, Shen W, He Y, et al.

·  Epithelial Sel1L is required for the maintenance of intestinal homeostasis[J]. Molecular biology of the cell, 2016, 27(3): 483-490， Sun S, Lourie R, Cohen S B, et al.

·  Effect of Sodium Dodecyl Sulfate Concentration on Supramolecular Gel Electrophoresis[J]. ChemNanoMat, 2016，Tazawa S, Kobayashi K, Yamanaka M.

·  Intergenic VNTR Polymorphism Upstream of rocA Alters Toxin Production and Enhances Virulence in Streptococcus pyogenes[J]. Infection and immunity, 2016: IAI. 00258-16，Zhu L, Olsen R J, Horstmann N, et al.

·  Ajuba Phosphorylation by CDK1 Promotes Cell Proliferation and Tumorigenesis[J]. Journal of Biological Chemistry, 2016: jbc. M116. 722751，Chen X, Stauffer S, Chen Y, et al.

·  Editorial: International Plant Proteomics Organization (INPPO) World Congress 2014[J]. Frontiers in Plant Science, 2016, 7，Heazlewood J L, Jorrín-Novo J V, Agrawal G K, et al.

·  Phosphoinositide kinase signaling controls ER-PM cross-talk[J]. Molecular biology of the cell, 2016, 27(7): 1170-1180，Omnus D J, Manford A G, Bader J M, et al.

·  A multiple covalent crosslinked soft hydrogel for bioseparation[J]. Chemical Communications, 2016, 52(15): 3247-3250，Liu Z, Fan L, Xiao H, et al.

·  Advances in crop proteomics: PTMs of proteins under abiotic stress[J]. Proteomics, 2016, 16(5): 847-865，Wu X, Gong F, Cao D, et al.

·  Cyclin-Dependent Kinase Co-Ordinates Carbohydrate Metabolism and Cell Cycle in S. cerevisiae[J]. Molecular cell, 2016, 62(4): 546-557，Zhao G, Chen Y, Carey L, et al.

·  Carbon Monoxide Gas Is Not Inert, but Global, in Its Consequences for Bacterial Gene Expression, Iron Acquisition, and Antibiotic Resistance[J]. Antioxidants & redox signaling, 2016，Wareham L K, Begg R, Jesse H E, et al.

·  Two-layer regulation of PAQR3 on ATG14-linked class III PtdIns3K activation upon glucose starvation[J]. Autophagy, 2016: 1-2，Xu D, Wang Z, Chen Y.

·  Regulation of sphingolipid biosynthesis by the morphogenesis checkpoint kinase Swe1[J]. Journal of Biological Chemistry, 2016, 291(5): 2524-2534，Chauhan N, Han G, Somashekarappa N, et al.

·  PAX5 tyrosine phosphorylation by SYK co-operatively functions with its serine phosphorylation to cancel the PAX5-dependent repression of BLIMP1: A mechanism for antigen-triggered plasma cell differentiation[J]. Biochemical and biophysical research communications, 2016, 475(2): 176-181，Inagaki Y, Hayakawa F, Hirano D, et al.

·  A Combined Computational and Genetic Approach Uncovers Network Interactions of the Cyanobacterial Circadian Clock[J]. Journal of Bacteriology, 2016: JB. 00235-16，Boyd J S, Cheng R R, Paddock M L, et al.

·  HuR mediates motility of human bone marrow-derived mesenchymal stem cells triggered by sphingosine 1-phosphate in liver fibrosis[J]. Journal of Molecular Medicine, 2016: 1-14，Chang N, Ge J, Xiu L, et al.

·  Combined replacement effects of human modified β-hexosaminidase B and GM2 activator protein on GM2 gangliosidoses fibroblasts[J]. Biochemistry and Biophysics Reports, 2016，Kitakaze K, Tasaki C, Tajima Y, et al.

·  Roseotoxin B Improves Allergic Contact Dermatitis through a Unique Anti-inflammatory Mechanism Involving Excessive Activation of Autophagy in Activated T-Lymphocytes[J]. Journal of Investigative Dermatology, 2016，Wang X, Hu C, Wu X, et al.

References on Phos-tag™ Chemistry

• Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry of phosphorylated compounds using a novel phosphate capture molecule, Rapid Communications of Mass Spectrometry, 17, 2075-2081 (2003), H. Takeda, A. Kawasaki, M. Takahashi, A. Yamada, and T. Koike 

• Recognition of phosphate monoester dianion by an alkoxide-bridged dinuclear zinc (II) complex, Dalton Transactions, 1189-1193 (2004), E. Kinoshita, M. Takahashi, H. Takeda, M. Shiro, and T. Koike

• Quantitative analysis of lysophosphatidic acid by time-of-flight mass spectrometry using a phosphate capture molecule, Journal of Lipid Research, 45, 2145-2150 (2004), T. Tanaka, H. Tsutsui, K. Hirano, T. Koike, A. Tokumura, and K. Satouchi

•  Production of 1,2-Didocosahexaenoyl Phosphatidylcholine by Bonito Muscle Lysophosphatidylcholine/Transacylase, Journal of Biochemistry,136, 477-483 (2004), K. Hirano, H. Matsui, T. Tanaka, F. Matsuura, K. Satouchi, and T. Koike

• Novel immobilized zinc(II) affinity chromatography for phosphopeptides and phosphorylated proteins, Journal of Separation Science, 28, 155-162 (2005), E. Kinoshita, A. Yamada, H. Takeda, E. Kinoshita-Kikuta, and T. Koike

• Detection and Quantification of On-Chip Phosphorylated Peptides by Surface Plasmon Resonance Imaging Techniques Using a Phosphate Capture Molecule, Analytical Chemistry, 77, 3979-3985 (2005), K. Inamori, M. Kyo, Y. Nishiya, Y. Inoue, T. Sonoda, E. Kinoshita, T. Koike, and Y. Katayama

• Phosphate-binding tag: A new tool to visualize phosphorylated proteins, Molecular & Cellular Proteomics, 5, 749-757 (2006), E. Kinoshita, E. Kinoshita-Kikuta, K. Takiyama, and T. Koike

• Enrichment of phosphorylated proteins from cell lysate using phosphate-affinity chromatography at physiological pH, Proteomics, 6, 5088-5095 (2006), E. Kinoshita-Kikuta, E. Kinoshita, A. Yamada, M. Endo, and T. Koike

• Separation of a phosphorylated histidine protein using phosphate affinity polyacrylamide gel electrophoresis, Analytical Biochemistry, 360, 160-162 (2007), S. Yamada, H. Nakamura, E. Kinoshita, E. Kinoshita-Kikuta, T. Koike, and Y. Shiro

• Label-free kinase profiling using phosphate-affinity polyacrylamide gel electrophresis, Molecular & Cellular Proteomics, 6, 356-366 (2007), E. Kinoshita-Kikuta, Y. Aoki, E. Kinoshita, and T. Koike

• A SNP genotyping method using phosphate-affinity polyacrylamide gel electrophoresis, Analytical Biochemistry, 361, 294-298 (2007), E. Kinoshita, E. Kinoshita-Kikuta, and T. Koike (The phosphate group at DNA-terminal is efficiently captured by Zn²⁺.Phos-tag.)

• Identification on Membrane and Characterization of Phosphoproteins Using an Alkoxide-Bridged Dinuclear Metal Complex as a Phosphate-Binding Tag Molecule, Journal of Biomolecular Techniques, 18, 278-286 (2007), T. Nakanishi, E. Ando, M. Furuta, E. Kinoshita, E. Kikuta-Kinoshita, T. Koike, S. Tsunasawa, and O. Nishimura

• A mobility shift detection method for DNA methylation analysis using phosphate affinity polyacrylamide gel electrophoresis, Analytical Biochemistry, 378, 102-104 (2008), E. Kinoshita-Kikuta, E. Kinoshita, and T. Koike

• Separation of phosphoprotein isotypes having the same number of phosphate groups using phosphate- affinity SDS-PAGE, Proteomics, 8, 2994-3003 (2008), E. Kinoshita, E. Kinoshita-Kikuta, M. Matsubara, S. Yamada, H. Nakamura, Y. Shiro, Y. Aoki, K. Okita, and T. Koike

• FANCI phosphorylation functions as a molecular switch to turn on the Fanconi anemia pathway, Nature Structural & Molecular Biology, 15, 1138-1146 (2008), M. Ishiai, H. Kitao, A. Smogorzewska, J. Tomida, A. Kinomura, E. Uchida, A. Saberi, E. Kinoshita, E. Kinoshita-Kikuta, T. Koike, S. Tashiro, S. J. Elledge, and M. Takata

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• Two-dimensional phosphate affinity gel electrophoresis for the analysis of phosphoprotein isotypes , Electrophoresis, 30, 550-559 (2009), E. Kinoshita, E. Kinoshita-Kikuta, M. Matsubara, Y. Aoki, S. Ohie, Y. Mouri, and T. Koike

• Formation of lysophosphatidic acid, a wound-healing lipid, during digestion of cabbage leaves, Bioscience, Biotechnology, and Biochemistry,73, 1293-300 (2009), T. Tanaka, G. Horiuchi, M. Matsuoka, K. Hirano, A. Tokumura, T. Koike, and K. Satouchi

• A Phos-tag-based fluorescence resonance energy transfer system for the analysis of the dephosphorylation of phosphopeptides, Analytical Biochemistry, 388, 235-241, (2009), K. Takiyama, E. Kinoshita, E. Kinoshita-Kikuta, Y. Fujioka, Y. Kubo, and T. Koike

• Phos-tag beads as an immunoblotting enhancer for selective detection of phosphoproteins in cell lysates, Analytical Biochemistry, 389, 83-85, (2009), E. Kinoshita-Kikuta, E. Kinoshita, and T. Koike

• Mobility shift detection of phosphorylation on large proteins using a Phos-tag SDS-PAGE gel strengthened with agarose, Proteomics, 9, 4098- 4101 (2009), E. Kinoshita, E. Kinoshita-Kikuta, H. Ujihara, and T. Koike

• Separation and detection of large phosphoproteins using Phos-tag SDS-PAGE, Nature Protocols, 4, 1513-1521 (2009), E. Kinoshita, E. Kinoshita-Kikuta, and T. Koike

• A clean-up technology for the simultaneous determination of lysophosphatidic acid and sphingosine-1-phosphate by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry using a phosphate-capture molecule, Phos-tag, Rapid Communications in Mass Spectrometry, 24, 1075-1084 (2010), J. Morishige, M. Urikura, H. Takagi, K. Hirano, T. Koike, T. Tanaka, and K. Satouchi

• Genotyping and mapping assay of single-nucleotide polymorphisms in CYP3A5 using DNA-binding zinc(II) complexes, Clinical Biochemistry, 43, 302-306 (2010), E. Kinoshita, E. Kinoshita-Kikuta, H. Nakashima, and T. Koike

• The DNA-binding activity of mouse DNA methyltransferase 1 is ragulated phosphorylation with casein kinase 1σ/ε, Biochemical Journal, 427, 489-497 (2010), Y. Sugiyama, N. Hatano, N. Sueyoshi, I. Suetake, S. Tajima, E. Kinoshita, E. Kinoshita-Kikuta, T. Koike, and I. Kameshita

  
  

ISOGEN II

总RNA及小RNA提取试剂

    离心后的沉淀              ISOGEN II 100 mL

　　ISOGEN II是从动物组织和培养细胞中提取总RNA和小RNA的试剂。本品是含有苯酚和胍的均匀液体，根据与细胞成分的相互作用，能一步分离RNA。

　　使用旧产品（ISOGEN和ISOGEN-LS）需要添加氯仿，要进行液相分离，而本产品则无需此步骤。在样品中加入ISOGEN II溶解或是均质之后，添加水， DNA、蛋白质、多糖类等沉淀（不溶化），进行离心分离后可以去除（右图）。用酒精沉淀、洗净、溶解上清液之后，能分离出高纯度的RNA。若配合ISOGENOME 使用，也能够跟ISOGEN一样从同一样本中提取DNA。

◆特点

　　•　无需使用氯仿分离RNA

　　•　相比旧产品（ISOGEN等），小RNA的提取率更高

　　•　可以把高分子RNA（＞200bases）和小RNA（＜200bases〉分离（也有不分离的方案）

　　•　混入的DNA较少，提取的RNA可以直接使用RT-PCR和定量RT-PCR

　　•　约一小时内可提取出RNA

◆用途

　　从活体组织提取RNA　等等

◆产品列表

运输方法

　　本品在室温下运输。产品到达后，在2~10℃条件下保存即可正常使用。

◆使用案例

操作流程

・除了本产品，根据需准备RNase free water、乙醇、4-溴甲氧基苯异丙醇。
・可使用聚丙烯管，使用前确认是否能够承受离心强度（12K x g）及ISOGEN II（苯酚）。
・离心操作建议在4-28℃进行。

实验数据

　　1.提取的RNA的电泳

　　2.检验出miRNA
　　3.RT-PCR
　　4.RNA提取共沉淀剂效果（测定吸光度）
　　5.RNA取量的基准
　　6.使用ISOGEN II从植物提取总RNA
　　7.使用ISOGEN II从植物提取高分子RNA及小RNA分离＜改变流程＞

Data 1：提取的RNA的电泳

　　按照ISOGEN及ISOGEN Ⅱ的操作顺序，提取RNA，进行电泳。

琼脂糖凝胶电泳
	M: RNA Ladder (编号:311-06261)   道1：ISOGEN    提取 总RNA (2μg) 道2：ISOGENⅡ 提取 总RNA (2μg) 道3：ISOGENⅡ 提取 高分子RNA (2μg) 道4：ISOGENⅡ 提取 小RNA (0.2μg)   1.2% Agarose S （甲醛变性） EtBr 染色

聚丙烯酰胺凝胶电泳
	道1：ISOGEN     提取 总RNA (5μg) 道2：ISOGENⅡ 提取 总RNA (5μg) 道3：ISOGENⅡ 提取 高分子RNA (5μg) 道4：ISOGENⅡ 提取 小RNA (0.5μg)   12% 聚丙烯酰胺凝胶 （Urea 变性） EtBr 染色

　　▶　道1、2中，ISOGENⅡ与ISOGEN一样，可提取全DNA。

　　▶　道3、4中，ISOGENⅡ可区分200bases以上的高分子RNA及10~200bases的小RNA。

Data 2：检验miRNA

对使用ISOGEN 及ISOGEN Ⅱ提取的总RNA进行3D-Gene Human miRNA Oligo chip分析 　▶　ISOGEN Ⅱ比ISOGEN更能显示miRNA。

Data 3：RT-PCR

　　使用ISOGEN 及ISOGEN Ⅱ从Hela细胞提RNA作为模板。通过RT-PCR检测出β-actin基因。同时，进行不产生RT反应的控制组实验。

PCR条件

1：模板RNA量 500ng 2：模板RNA量 100ng 3：模板RNA量 20ng M：Gene Ladder  （编号：316-06951）

　　▶　使用ISOGEN Ⅱ提取的RNA比使用ISOGEN抽提的RNA混入DNA较少。

Data 4：RNA提取共沉淀剂效果（测定吸光度）

　　ISOGEN Ⅱ的使用指南中分离小RNA时，对使用共沉淀剂Ethachinmate（Nippon Gene 产品编号：312-01791）或者Glycogen（和光纯药工业 产品编号：077-05311）时提取到小RNA情况进行了比较。

方法   按照使用指南，从Hela细胞及小鼠肝脏回收小RNA，检测吸光度，计算小RNA的含量。将使用Ethachinmate提取到的数量看作1，在图表中列出了含量的相对值。

　　▶　使用Glycogen作为共沉淀剂回收小RNA时的回收量较多，变异较少。

Data 5：RNA取量的基准

Data 6：使用ISOGEN II从植物提取总RNA

　　从植物抽提全RNA例子 (PDF 222KB)

Data 7：使用ISOGEN II从植物抽提高分子RNA及小RNA分离＜变化流程＞

　　从植物抽提高分子RNA及小RNA离析例子 (PDF 218KB)

◆注意事项

　　•　本品是实验研究用的试剂，不能作为医药品或其他目的使用。另外，如果您对试剂的基本的知识不了解，请不要使用。

　　•　ISOGEN II是非医疗用剧毒物（苯酚制剂），请小心操作。

　　•　使用时请穿着适当的防护设备（手套、眼镜等）。

　　•　操作场所保持通风，以免吸入蒸汽。

　　•　如果接触皮肤或眼睛，请用大量的水冲洗15分钟或以上，并及时就诊。

　　•　请按照说明书进行操作。

　　•　对于因不按照说明书操作而导致的事故，Nippon Gene不负任何责任。

◆备注

　　本产品是实验研究用试剂，请勿用作医疗用途。

◆资料 Data Sheet

产品说明书

　　•　ISOGEN II　产品说明书第三版（100 ml产品小冊子） (PDF 195KB)

　　•　(内容与册子相同） (PDF 168KB)

SDS(Safety Data Sheet)

　　•　ISOGEN II 　安全数据图表(PDF)

宣传单

　　•　册子（PDF 2MB）

◆相关产品

　　 •　RNA抽提用试剂　ISOGEN、ISOGEN-LS

　　 •　RNA抽提用试剂　ISOGEN with Spin Column

　　 •　酒精沉淀用共沉淀剂　Ethachinmate

　　 •　缓冲器　ddWater, TE（pH 8.0）

Q:　　可以从全血等血液样本中抽提RNA吗？

A:　　ISOGEN II难以提取出高纯度的RNA，建议使用ISOGEN-LS。

Q:　　可以提取DNA或蛋白质吗？

A:　　旧法使用ISOGEN，可以从同一样本回收DNA及蛋白质，但由于此次研发的是提取RNA的方法，ISOGEN II中混入的DNA较少，无法提取DNA及蛋白质。不过，配合使用DNA提取试剂ISOGENOME，可回收DNA样本。

　　[ 1 ]　Chomczynski, P. ： Reagents and methods for isolation of purified RNA, US and International Patents Pending.

　　[ 2 ]　Chomczynski, P. and Sacchi, N.：”Single step method of RNA isolation by acid guanidinium thiocyanate-phenol- chloroform extraction”, Anal. Biochem，162, 156-159（1987）

　　[ 3 ]　Chomczynski, P.：”A reagent for the single-step simultaneous isolation of RNA, DNA and proteins from cell and tissue samples”, Biotechniques,15, 532-537（1993）

　　[ 4 ]　Wilfinger, W., Mackey, K. and Chomczynski, P.：”Effect of pH and ionic strength on the spectrophotometric assessment of nucleic acid purity”,Biotechniques, 22, 474-481（1997）

Ethachinmate共沉淀剂

Ethachinmate

◆原理

       共沉淀剂Ethachinmate是基因工程学的标准时机。作为畅销产品，效果显著。
       Ethachinmate适用于核酸（DNA与RNA）乙醇或isopropanol沉淀时的丙烯酰胺系高分子载体溶液，在盐类存在的情况下（例如 ＞0.1mol/L Sodium Acetate）加上Ethachinmate进行酒精沉淀，能得到如下优点。

◆优点・特色

● 可回收微量核酸
   可定量性回收20ng/mL以上的DNA（＞100个碱基对）及（＞120个碱基）。
● 乙醇沉淀迅速
   无需-20℃或-80℃孵化，添加乙醇后可直接离心。
● 酶反应无障碍
   回收的核酸，在水及缓冲液中容易溶解。混在一起的Ethachinmate完全不阻碍酶反应。
● 沉淀可观察到
   因为Ethachinmate自己根据究竟沉淀形成沉淀，无需担心微量核酸冲洗了重要样品。

无DNase，RNase活性测试剂

用一下DNase及RNase free方法可确认是否含有Ethachinmate

DNase实验：向1μgλ/Hin d III 中加入3μLEthachinmate，在37℃的恒温箱中放置16小时候，进行琼脂糖凝胶电泳，图像确认无变化。

RNase实验：向2μg 16S及23S的rRNA中加入3μL Ethachinmate，在37℃的恒温箱中放置16小时后，进行琼脂糖凝胶电泳，图像确认无变化。

◆使用方法

DNA或者RNA溶液（100μL）

↓←3 mol/L Sodium Acetate（添加），3,3μL*1）

↓←Ethachinmate 1μL*2）

↓←涡流

↓←2~2.5倍量的乙醇

↓涡流*3）

↓10kgxg室温下反应5分钟*4）

沉淀*5）

*1）（氯）盐的最终浓度必须在0.1mol/L以上

*2）每100μL DNA或RNA溶液中加入1μL  Ethachinmate。但溶液量为300μL以上时，加入3μL可达到饱和。

*2）而且再次进行上述操作时，无需添加Ethachinamte。　（若反复添加Ethachinmate会导致DNA溶液变

*2）得黏稠，影响到后面的操作。）

*3）进行充分混匀，可实现微量DNA的定量回收。

*4）不必进行冷却

*5）沉淀可目测。溶解于其他缓冲液等中，可直接作为各种酶底物使用。而且，必须用70%的乙醇进行洗涤。

Q：    有没有DNase，Rnase杂质？
A：    没有。Ethachinmate用以下的方法可以确认无DNae、RNase活性。
          Dnase 实验：向1μg的/Hin d III中加入3μL Ethachinmate，在37℃的恒温箱中放置16小时候，进行琼脂

          糖凝胶电泳，图像确认无变化。
          RNase实验：向2μg 16S及23S的rRNA中加入3μL Ethachinmate，在37℃的恒温箱中放置16小时后，进

          行琼脂糖凝胶电泳，图像确认无变化。

Q：    可以使用RNA回收吗？
A：    没有问题。

Q：    高效回收的核酸浓度和核酸链长度是多少？
A：    可回收几乎所有的20ng/ml 以上的DNA（100bp以上）及RNA（120 base以上）。

Q：    260nm的吸光度会影响定量吗？
A：    无影响。

Q：    100μL DNA溶液对应加入1μL Ethachinmate。若DNA溶液少于100μL，应该怎样做才好？
A：    请加入1μL Ethachinmate。为了DNA沉淀可目测，需要1μL以上的Ethachinmate。另外，3M Sodium

          Acetate及1μL的Ethachinmate。

Q：    300μL以上的DNA溶液，Ethachinmate的添加量是多少？
A：    DNA溶液为300μL以上，无论DNA溶液的量是多少，都请加入3μL Ethachinmate。无需加入过量的Eth-

          achinmate。另外，3M Sodium Acetate请按此比例换算适量添加。
          例：DNA溶液为600μL，则加入19.8μL的3M Sodium Acetate及3μL的Ethachinmate

Q：    DNA溶液进行2次以上乙醇沉淀时，最好第二次以后也每次加上Ethachinmate吗？
A：    Ethachinmate添加一次无需再添加，如果重复添加Ethachinmate，DNA溶液会变得黏稠，将阻碍以后的

          操作。

Q：    冷冻时，会不会影响Ethachinmate的效果？
A：    不会

Q：    高压处理会不会影响Ethachinmate的效果？
A：    不会

Q：    如果用苯酚/氯仿处理含Ethachinmate的DNA溶液，会不会影响Ethachinmate的效果？
A：    不会

Q：    对电泳模型有没有影响？
A：    没有，但根据电泳条件的不同，数十kbp以上的DNA会产生宽频带条。

Q：    对限制性内切酶反应有没有影响？
A：    没有。

Q：    对T4 DNA Ligase反应有没有影响？
A：    没有。

Q：    对AMV Reverse Transcroptase的cDNA合成反应有没有影响？
A：    没有。

Q：    对使用Taq DNA Polymerase的PCR反应有没有影响？
A：    没有。

Q：    对Klenow Flagment反应有没有影响？
A：    没有。

Q：    对大肠杆菌的转化有没有影响？
A：    没有。

Q：    对in vitro组装有没有影响？
A：    λ 噬菌体的in vitro组装效率稍微下降。

Q：    加入Ethachinmate，乙醇沉淀会形成核苷酸沉淀吗？
A：    使用8mer及17mer的的低聚核苷酸进行实验，与是否添加Ethachinmate无关，回收率没有变化。

Q：    在含有甲酰胺的杂交溶液中有无变性吗？
A：    无变性。

Q：    对blotting有没有影响？
A：    没有。

Q：    对序列反应有没有影响？
A：    不影响使用ABI Prism 377的周序列反应。对脱氧发的序列反应也没有影响。

Q：    添加Ethachinmate是否有助于从管中脱落沉淀？
A：    要看管的材质。比如Eppendorf公司制的Safe-Lock管有容易脱落的倾向。