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查看更多产品信息 Custom RNA, Stealth, HPLC - FAQs (100001863)
26 个常见问题解答
我们提供下列体内siRNA对照：
•Ambion In Vivo阴性对照 #1 siRNA（货号4457287, 5nmol; 4459405, 50 nmol; 4457289, 250 nmol）
•Ambion In Vivo GAPDH阳性对照siRNA（货号4457288, 5 nmol; 4459407, 50 nmol, 4457291, 250 nmol）
•Ambion In Vivo Factor VII阳性对照siRNA（货号4459408, 50 nmol; 4457292, 250 nmol）
•Silencer Select阴性对照1号 siRNA, in vivo ready（货号4404020, 250 nmol）
•Silencer GAPDH阳性对照siRNA, in vivo ready（货号4404025, 250 nmol）
Stealth siRNA比非修饰siRNA在血清中稳定的多。这一高稳定性对于体内实验尤其有好处，因为在体内RNAi分子会暴露于更多蛋白酶。通过与 Intradigm合作，Stealth RNAi进行瘤内注射时已被证明具有体内活性。
用于体内实验的Stealth siRNA双链可以使用BLOCK-iT RNAi Express订购，而Ambion in vivo siRNA可以使用此工具（https://www.thermofisher.com/us/en/home/life-science/rnai/introduction-to-in-vivo-rnai/ambion-in-vivo-sirna.html?icid=cvc-invivo-sirna-c3t1）订购。
推荐进行内毒素检测以减少引发免疫反应的风险。
请联系rnairesearcher@invitrogen.com咨询大规模体内siRNA订购或填写询价表格（ www.thermofisher.com/us/en/home/global/forms/custom-sirna-mirna-quote-request.html）。
有数种不同的方法可以用于siRNA导入，包括多种局部导入技术和系统导入技术。我们提供一种脂质体试剂Invivofectamine 3.0用于体内siRNA系统导入。点击此处（https://www.thermofisher.com/us/en/home/life-science/rnai/introduction-to-in-vivo-rnai/invivofectamine-reagent/invivofectamine-reagent-data.html）了解更多关于此试剂的信息。
RNAi可以使用两种不同方法导入——合成的siRNA双链或通过质粒或病毒载体表达的siRNA（shRNA，miRNAi）。siRNA正成为快速发展的治疗应用的首选方法。它们易于使用，易于设计，并且易于合成。siRNA可以被快速鉴定并且可以同时靶向多个基因。对于RNAi载体而言，其表达可能由于质粒在基因组的稳定整合而更稳定持久，而它们也可以靶向非分裂细胞例如干细胞、淋巴细胞和神经元。RNAi载体的不足是插入突变导致的致癌风险，以及长期基因沉默和/或细胞内大量siRNA的存在导致的不可预测的毒性（Grimm D. et al.: Nature 441: 537-541 (2006)）。
对于 Stealth siRNA我们推荐选用HPLC in vivo纯化级别。对于Ambion In Vivo siRNA，我们推荐in vivo ready纯化级别。
我们已经证明标记的Stealth和Silencer siRNA不会影响它们的基因敲低能力。一个替代的方法是可以将非标记siRNA的和标记的对照siRNA混合；该方法更多用于体内siRNA实验，并可以使临床研究得以推进而不受标记可能带来的效应的影响。
化学修饰的siRNA双链相比标准siRNA双链有很多优势，包括低脱靶效应，高稳定性，以及低毒性。因为这些原因，我们推荐使用化学修饰的siRNA用于体内实验。对于一些目的只是确定生物分布的探索实验，非修饰的siRNA和荧光对照会更有帮助并且稍微便宜一些。
请提供基因数据库编号或核酸序列。您还可以选择您需要靶向的目的基因的特定区域：ORF, 5’ UTR或3’ UTR以及物种。siRNA的GC含量也可以选择。最后，您可以将您的输入序列在数据库中搜索比对以确保siRNA靶向特异性序列。
您可以订购Block-it RNA寡核苷酸或定制RNA寡核苷酸。Block-it RNA寡核苷酸是21-23碱基、非修饰的序列特异性RNA双链。要设计和订购BLOCK-iT siRNA，请访问BLOCK-iT RNAi Designer（https://rnaidesigner.thermofisher.com/rnaiexpress/）。定制siRNA也可以通过电子邮件订购（https://www.thermofisher.com/us/en/home/life-science/rnai/rnai-email-order-forms.html）。
定制RNA寡核苷酸可以以单链或双链RNA订购。这些寡核苷酸最长50 bp，可以选择5’生物素、荧光素或磷酸基团修饰。它们仅能以脱盐纯化方式订购。要订购定制RNA寡核苷酸，请访问该网页（https://www.thermofisher.com/order/catalog/en/US/adirect/lt?cmd=ConfigureRNASingleTube）。
如果需要体内级别的和/或5’正义链标记（Biotin, Alexa Fluor 488, 546, 555, 647, 680, 750），您可以在我们的RNAi in vivo Designer页面（https://rnaidesigner.thermofisher.com/rnaiexpress/inVivoService.jsp?icid=fr-rnaiordering-3）订购。这些siRNA可以以HLPC纯化级别或体内脱盐级别或体内HPLC级别进行订购。
We offer the following in vivo siRNA controls:
Invitrogen In Vivo Negative Control #1 siRNA (Cat. No. 4457287, 5nmol; 4459405, 50 nmol; 4457289, 250 nmol)
Invitrogen In Vivo GAPDH Positive control siRNA (Cat. No. 4457288, 5 nmol; 4459407, 50 nmol, 4457291, 250 nmol)
Invitrogen In Vivo Factor VII Positive control siRNA (Cat. No. 4459408, 50 nmol; 4457292, 250 nmol)
Silencer Select Negative Control No. 1 siRNA, in vivo ready (Cat. No. 4404020, 250 nmol)
Silencer GAPDH Positive Control siRNA, in vivo ready (Cat. No. 4404025, 250 nmol)
Find additional tips, troubleshooting help, and resources within our RNAi Support Center.
Stealth siRNA is markedly more stable in serum than unmodified siRNA. This increased stability is particularly beneficial in vivo where the RNAi may be exposed to more proteases. In collaboration with Intradigm, Stealth RNAi has been demonstrated to have activity in vivo when injected intra-tumorally.
Find additional tips, troubleshooting help, and resources within our RNAi Support Center.
Pre-designed Stealth RNAi and Ambion In Vivo siRNA can be ordered directly from our pre-designed siRNA search interface (https://www.thermofisher.com/us/en/home/life-science/rnai/introduction-to-in-vivo-rnai.html?open=faq#search) to target human, mouse, or rat genes.
Custom siRNA designs for in vivo experiments can be ordered from two interfaces, depending on the product format being requested: Custom Ambion In Vivo siRNA is ordered from the GeneAssist Custom siRNA Builder (https://www.thermofisher.com/order/custom-genomic-products/tools/sirna/), while Stealth RNAi is ordered using the BLOCK-iT RNAi Express for in vivo synthetics (https://rnaidesigner.thermofisher.com/rnaiexpress/inVivoService.jsp).
Find additional tips, troubleshooting help, and resources within our RNAi Support Center.
It is recommended to reduce the chance of eliciting an immune response.
Find additional tips, troubleshooting help, and resources within our RNAi Support Center.
Please contact rnairesearcher@invitrogen.com regarding large scale in vivo siRNA or fill out the quote request form (www.thermofisher.com/us/en/home/global/forms/custom-sirna-mirna-quote-request.html).
Find additional tips, troubleshooting help, and resources within our RNAi Support Center.
Several different approaches have been used for siRNA delivery, including various local delivery techniques and systemic delivery. We offer a lipid reagent, Invivofectamine 3.0 Transfection Reagent for in vivo siRNA systemic delivery. Learn more about this reagent here (https://www.thermofisher.com/us/en/home/life-science/rnai/introduction-to-in-vivo-rnai/invivofectamine-reagent.html).
Find additional tips, troubleshooting help, and resources within our RNAi Support Center.
RNAi can be delivered using two different approaches-siRNA synthetic duplexes or siRNA expressed from plasmids or viral vectors (shRNA, miRNAi). siRNA are becoming the method of choice for the fast development of therapeutics. They are easy to use, easy to design, and easy to synthesize. siRNA can be rapidly identified and multiple genes can be targeted at the same time. With RNAi vectors, the expression will be steadier as a result of the possibility of stable integration of the plasmid into the genome, and they have the ability to target nondividing cells such as stem cells, lymphocytes and neurons. The drawbacks are the danger of oncogenic transformation from insertional mutagenesis, and unanticipated toxicity from long-term silencing of human genes and/or having high amounts of siRNA inside the cell (Grimm D. et al.: Nature 441: 537-541 (2006)).
Find additional tips, troubleshooting help, and resources within our RNAi Support Center.
For Stealth siRNA we recommend to select HPLC in vivo purity. For Invitrogen In Vivo siRNA, we recommend in vivo ready purity.
Find additional tips, troubleshooting help, and resources within our RNAi Support Center.
We have demonstrated that labeling Stealth and Silencer siRNA does not hamper their knockdown potency. An alternative approach is to mix unlabeled duplexes with labeled control duplexes; this method is more commonly used with in vivo siRNA, and allows progression to clinical research unhindered by questions about the possible effects of a label.
Find additional tips, troubleshooting help, and resources within our RNAi Support Center.
Chemically modified siRNA duplexes have a number of advantages over standard siRNA duplexes, including the minimization of off-target effects, enhanced stability, and reduced toxicity. For these reasons, chemically modified siRNA duplexes are recommended for in vivo siRNA experiments. For some pilot experiments, where the only aim is to determine biodistribution, unmodified siRNA and fluorescent controls are useful and slightly more cost-effective.
Find additional tips, troubleshooting help, and resources within our RNAi Support Center.
Please provide either the accession number or nucleotide sequence. You will also have the choice of the region of the gene you would like to target: ORF, 5' UTR, or 3' UTR and the species. The percentage G/C content of the siRNA can also be chosen. Lastly, you can have your input sequence blasted against sequences in the database to ensure unique regions are used to design the siRNAs.
Find additional tips, troubleshooting help, and resources within our RNAi Support Center.
You could order a Block-it RNA oligo or custom RNA oligo. Block-it RNA oligos are small 21-23 unmodified, nucleotide sequence-specific RNA duplexes. To design and order BLOCK-iT siRNAs, please visit the BLOCK-iT RNAi Designer (http://rnaidesigner.thermofisher.com/rnaiexpress/). Custom siRNAs can also be ordered by email (http://www.thermofisher.com/us/en/home/life-science/rnai/rnai-email-order-forms.html).
Custom RNA oligos can be ordered as single stranded oligos or duplexed to create a double stranded RNA. The oligo can be up to 50 bp in length, with the option of a biotin, fluorescein or phosphate 5' modification. These oligos will be delivered at desalted purity only. To order custom RNA oligos, visit this website (https://www.thermofisher.com/order/catalog/en/US/adirect/lt?cmd=ConfigureRNASingleTube).
Find additional tips, troubleshooting help, and resources within our RNAi Support Center.
If in vivo purity and/or 5' sense label (Biotin, Alexa Fluor 488, 546, 555, 647, 680, 750) is required, you can order the siRNA through our RNAi in vivo Designer page (http://rnaidesigner.thermofisher.com/rnaiexpress/inVivoService.jsp). These siRNA can be ordered as HLPC or desalted in vivo purity or HPLC in vivo purity.
Find additional tips, troubleshooting help, and resources within our RNAi Support Center.