AQUApure Linear Di-Ub Chains (M1-linked) Protein, CF Summary
Ubiquitin chains vary in length, linkage, and function. Linear (Met1)-linked Di-Ubiquitin Chains (Ub2) are ideal for investigating Ubiquitin-binding proteins and as substrates for Ubiquitin-specific isopeptidases. Reaction conditions will need to be optimized for each specific application. IMPORTANT: Heating this product in SDS-PAGE buffer or terminating reactions containing this product with heated SDS-PAGE buffer could lead to unexpected, high apparent molecular weight banding or smearing on gels that is not representative of product purity. For optimal results, we recommend incubation in SDS-PAGE buffer + DTT at <40 °c="" for="" 20="" minutes="" prior="" to="" gel="">
Product Datasheets
Carrier Free
CF stands for Carrier Free (CF). We typically add Bovine Serum Albumin (BSA) as a carrier protein to our recombinant proteins.Adding a carrier protein enhances protein stability, increases shelf-life, and allows the recombinant protein to be stored at a more dilute concentration.The carrier free version does not contain BSA.
In general, we advise purchasing the recombinant protein with BSA for use in cell or tissue culture, or as an ELISA standard.In contrast, the carrier free protein is recommended for applications, in which the presence of BSA could interfere.
UC-700B
| Formulation | 1 mg/ml (58 μM) in sterile, deionized water |
| Shipping | The product is shipped with polar packs. Upon receipt, store it immediately at the temperature recommended below. |
| Stability & Storage: | Use a manual defrost freezer and avoid repeated freeze-thaw cycles.
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Reconstitution Calculator
Background: Di-Ubiquitin
Linkage specific Poly-Ubiquitin chains may be used as a substrate for in vitro reactions with deubiquitinating enzymes ("DUB"s") that cleave the peptide or isopeptide linkage between adjacent Ubiquitin molecules. Poly-Ubiquitin chains can also be used to investigate mechanisms of binding and recognition between the chains and other proteins that contain Ubiquitin-Associated domains (UBAs), Ubiquitin-interacting motifs (UIMs), ZnF"s and/or other Ubiquitin-sensing elements.
Linear ("M1")-linked Di-Ubiquitin chains are manufactured using recombinant methods to avoid the potential for contaminating synthetic intermediates. The correctness of linkage and purity of each production lot is assessed using the Absolute Quantitation of Ubiquitin method (Ub-AQUA), an LCMS-based technique that provides extremely accurate information on the composition of Poly-Ubiquitin samples.
- Kirkpatrick D.S., et al. (2006) Nat Cell Biol. 8(7) : 700-10
- Ordureau, A., et al. (2014) Mol. Cell 56(3) : 360–375
- Ordureau, A., et al. (2015) Pro. Nat. Acad. of Sci. USA 112(21) : 6637–6642
- Phu L., et al. (2011) Mol Cell Proteomics 10(5) : M110.003756
Citations for AQUApure Linear Di-Ub Chains (M1-linked) Protein, CF
R&D Systems personnel manually curate a database that contains references using R&D Systems products.The data collected includes not only links to publications in PubMed,but also provides information about sample types, species, and experimental conditions.
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- Post-translational Modification of OTULIN Regulates Ubiquitin Dynamics and Cell DeathAuthors: T Douglas, M SalehCell Rep, 2019;29(11):3652-3663.e5.Species: HumanSample Types: ProteinApplications: Bioassay
- ZUFSP Deubiquitylates K63-Linked Polyubiquitin Chains to Promote Genome StabilityAuthors: P Haahr, N Borgermann, X Guo, D Typas, D Achuthanku, S Hoffmann, R Shearer, TK Sixma, N MailandMol. Cell, 2018;0(0):.Species: HumanSample Types: Cell LysatesApplications: Bioassay
- Cholesterol and fatty acids regulate cysteine ubiquitylation of ACAT2 through competitive oxidationAuthors: YJ Wang, Y Bian, J Luo, M Lu, Y Xiong, SY Guo, HY Yin, X Lin, Q Li, CCY Chang, TY Chang, BL Li, BL SongNat. Cell Biol., 2017;19(7):808-819.Species: N/ASample Types: Applications: Control
- Myosin VI Contains a Compact Structural Motif that Binds to Ubiquitin ChainsAuthors: F He, HP Wollscheid, U Nowicka, M Biancospin, E Valentini, A Ehlinger, F Acconcia, E Magistrati, S Polo, KJ WaltersCell Rep, 2016;14(11):2683-94.Species: HumanSample Types: Recombinant ProteinApplications: Bioassay
- USP45 deubiquitylase controls ERCC1-XPF endonuclease-mediated DNA damageresponses.Authors: Perez-Oliva A, Lachaud C, Szyniarowski P, Munoz I, Macartney T, Hickson I, Rouse J, Alessi DEMBO J, 2015;34(3):326-43.
- Deubiquitinase-based analysis of ubiquitin chain architecture using Ubiquitin Chain Restriction (UbiCRest).Authors: Hospenthal, Manuela, Mevissen, Tycho E, Komander, DavidNat Protoc, 2015;10(2):349-61.Species: HumanSample Types: Protein
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早期基因捕获载体的特点是当报告基因与内源基因的读码框一致时,产生由内源蛋白的N 末端与报告基因蛋白融合的活性蛋白质 。近来对这一类载体所做的改进是在报告基因与SA 序列间插入一来源于脑、心肌炎病毒的IRES ,这样即使报告基因与捕获基因不发生融合也可产生报告基因翻译产物。尽管ES 细胞中大量基因具转录活性,为了获得全基因组突变克隆必须使用不依赖于捕获基因表达的筛选策略。
我从未做过，不知道以下四种：
StandardcDNALibraries
LargeinsertcDNALibraries
NormalizedcDNALibraries
SubtractedcDNALibraries
其中StandardcDNALibraries和NormalizedcDNALibraries，SubtractedcDNALibraries有啥区别阿？
我只是想构建文库，以后可以用来做southern的probe，或者以后可以拿来做做蛋白方面的等等。
还有两个问题：
1.大家cDNA文库构建的时候都是先直接提取mRNA的吧？大家推荐以下用什么kit？
2.先提取总RNA然后再提取mRNA；直接从组织中提取mRNA.这两者有什么主要区别？
不好意思，如果提的问题是有错误，多多包涵。。。
tRNA不是一条直的单链，而是弯曲的，呈现一个三叶草的形状。在弯曲的部位，tRNA自己的碱基跟自己的碱基互补配对连起来，碱基对中存在氢键。
其他的RNA一般为单链不存在氢链，但双链的由于碱基互补配对存在氢键。
已经在mirbase数据库下载水稻的所有miRNA序列，可以比对某种昆虫的转录组文库寻找靶基因吗？
可以用targetscan或者miRwalk网站吗？有什么方法吗？
目标基因全长1800bp，通过重叠延伸的方式获得的自杀敲除组件，左右同源序列均为500bp左右，构建自杀载体（确认载体没有问题），期望进行目标基因的失活处理，但将近1个月的时间，不见任何目的克隆，故在此请各位战友指点。
采用电转的方式将自杀载体（约6700bp）导入黄色短杆菌，感受态的细胞制备采用相关文献方式，甘氨酸和吐温-30的方式进行摇菌并制备电转感受态，1800v电转（电压进行过梯度，1200、1500、1800、2200、2500。1800v效果相对较好，故选之），但电转效率仍是较低，复苏2h，浓缩涂板LBG+kan（kan浓度20ug/ml）,平板克隆很少，几次都只有20个左右的克隆。
1）克隆很少，原因可能跟菌株的电转效率有关，不知战友有没有谷氨酸棒杆菌或黄色短杆菌相关较好的电转经验，还请指教？
2）抗性平板克隆很少，还可能和重组效率有关，但根据pk18mobsacB质粒的特性，既然能在抗性平板生长，理论上应该是已经进行了一次重组了，但结果是不管我用pk18载体本身的Kan序列引物还是最终诱导进行PCR验证，均未证实到一次重组的发生，更不要说获得敲除失活的目的菌株了。问题是这个带有抗性的克隆到底是携带了什么使其同样具有抗性(显微检验不是杂菌）？如何有效提高或促进自杀载体在宿主内的重组效率呢？或是更有效的准确的检测验证手段？？
3）利用自杀载体的方式进行黄色短杆菌（或谷氨酸棒杆菌）的基因敲除缺失比较不易，不知有没有正在做这方面相关研究的战友，希望能够多多交流和学习。。。
不知道我的疑惑讲清楚了没有，若哪里不清楚还请指出，我再细说。。。期望xdjm们的帮助啊。。。先谢过了。。
在分子生物学中，探针是根据碱基互补的原理，用来与特定的DNA片段做杂交以对特定的DNA片段进行检测，如Southern杂交
引物是用来扩增DNA序列的，因为核苷酸必须要连接在3-OH上才能够合成延伸，引物就是用来提供3-OH的。
只要是符合要求的DNA片段都可以用来做探针和引物，引物是用来扩增DNA序列，探针用来与特定的DNA片段做分子杂交的
引物设计略有不一样，但都可用Primer5设计。
