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查看更多产品信息 GeneArt™ Gene Set-up Service Genes <3 kb (Service fee) - FAQs (878017DE)
40 个常见问题解答
试剂盒和定制服务有不同的优点。请点击此处（https://www.thermofisher.com/us/en/home/life-science/cloning/gene-synthesis/geneart-gene-synthesis-product-selection-guide.html）查看一份对技术，递送和生产时间进行比较的选择指南，以决定哪种最适合您。
我们专利化的GeneOptimizer软件（https://www.thermofisher.com/us/en/home/life-science/cloning/gene-synthesis/geneart-gene-synthesis/geneoptimizer.html）计算编码目的蛋白所需的最优DNA序列（基因优化）。根据表达物种的密码子偏好改变基因的密码子使用情况（密码子优化）仅是基因优化的一个方面。GeneOptimizer软件目前评估多达50个可能影响mRNA稳定性的因素，例如极端GC含量，核糖体结合位点，通用序列及潜在的剪接位点，重复序列，以及二级结构。mRNA分子稳定性的提高通常会提高蛋白产量。
GeneArt基因合成服务包括基因优化（如果要求的话），合成DNA，克隆至我们的标准载体，以及提供5 µg冻干的DNA和一张包含计算机分析和质量控制信息的压缩光碟。
是的。我们可以将您的基因亚克隆到您提供的载体内。您将收到将您的目的基因分别克隆到GeneArt克隆载体和您自己的载体的两个重组质粒各5 µg（以及两个载体的质量控制信息）。
不幸的是，这个载体不包含适用于蛋白表达的启动子或终止子等表达元件。我们建议通过限制性内切酶克隆将您的合成基因亚克隆到您选择的表达载体中，或者我们可以为您进行定制服务。
是的，您可以保护您不希望被GeneOptimizer过程优化的特殊区域或限制性酶切位点。在门户网站，在“Optimize sequence（优化序列）”之下，选择“Protect Restriction Sites（保护限制性酶切位点）”或“Protect Sequence Parts（保护序列部分）”。
其ori是ColE1。< / p >
由于我们的生产流程，我们不提供这种选项。限制性内切酶的引入通常会切去标准载体的大部分多克隆位点。< / p >
基因通常被克隆进pMx标准载体。这些载体包含：
•一个多克隆位点
•一个复制起始位点（colE1）
•一个抗生素抗性标记（氨苄青霉素，卡那霉素，链霉素，氯霉素）；请注意标准订单不保证任何特定的标准载体。
有多个因素需要考虑：CG含量，重复评分，直接重复，和二级结构，以及指定的宿主生物和基因的总长度。当判断一个基因或一个序列的一部分是否“复杂”时，所有这些因素都要考虑进去。
该超高速递送选项仅适用于非复杂基因，大小不超过3kb。如果你的基因不符合这些要求，这个选项就会变成灰色。
如果不满足此标准，可以将该基因做为新的基因合成项目订购。
目前，我们不提供这种服务。亚克隆可以使用基于限制性内切酶的载体或通过BP/LR克隆酶反应进入Gateway载体来实现。或者，我们提供的GeneArt Strings DNA片段可以直接被克隆到TOPO Zero Blunt载体中。
除另有要求外，您将获得5 µg 冻干质粒(合成的基因克隆到pMx质粒中)。收到后，可以重悬DNA，并分成小份保存在-20°C。您还将收到一张CD，里面有您的DNA测序比对数据。
由于微生物抗性标记的大小不同，其大小略有差异。pMA：2.5kb，pMS：2.6kb，pMC：2.3kb，pMK：2.4kb。
GeneOptimizer过程使用一种多参数的方法进行基因序列分析。它在进行基因序列优化时考虑密码子使用，GC含量，潜在剪接位点，直接重复序列，RNA二级结构，以及不稳定序列。
理论上，没有限制。但是，非常长的DNA片段（超过10 kb）需要克隆进一个低拷贝载体以确保遗传稳定性。因此，长度的提高意味着生产时间的延长。
我们提供GeneArt Strings DNA Fragments service，提供定制长度可达1000 bp的线性双链DNA片段。访问www.thermofisher.com，搜索“GeneArt Strings DNA Fragments”，了解更多关于该产品的信息。
有的，所有pMx载体包含一个M13 – 20引物结合位点和一个M13-R引物结合位点。其序列如下：
•M13 – 20引物结合位点：5'-TTGTAAAACGACGGCCAG-3'
•M13-R引物结合位点：5'-GGAAACAGCTATGACCAT-3'
pMx是用于克隆您感兴趣的合成基因的标准GeneArt克隆载体。& # 147;x # 148;代表抗生素的选择。因此，pMA代表氨苄西林，pMK代表卡那霉素，pMS代表壮观霉素, pMC代表氯霉素，pMZ代表Zeocin抗生素。所有载体均含有一个多克隆位点、一个ORI (colE1)和一个抗生素抗性标记，以及用于测序的M13-20/M13-R引物结合位点。
The kit and custom services offer different advantages. Please click here (http://www.thermofisher.com/us/en/home/life-science/cloning/gene-synthesis/geneart-gene-synthesis-product-selection-guide.html) to view a selection guide comparing the technologies, including deliverables and production time, to determine what is best for you.
Our proprietary GeneOptimizer software (https://www.thermofisher.com/us/en/home/life-science/cloning/gene-synthesis/geneart-gene-synthesis/geneoptimizer.html) calculates the optimal DNA sequence needed to encode the protein of interest (gene optimization). Adapting the codon usage of the gene to the codon preferences of the expression organism (codon optimization) is just one of the parameters addressed. The GeneOptimizer software currently evaluates several additional parameters that can compromise mRNA stability, such as extreme GC content, ribosomal binding sites, consensus and cryptic splice sites, repeats, and secondary structures. Increased numbers of stable mRNA molecules often lead to higher yields of protein.
GeneArt Gene Synthesis Service includes gene optimization (if requested), synthesis of the DNA, cloning into our standard vector, and delivery of 5 µg of lyophilized DNA plus a compact disc containing the in silico analysis and quality control information.
Yes. We can subclone your gene into a vector you provide. You will receive 5 µg each of your gene in a GeneArt cloning vector and in your vector (plus the quality control information for both constructs).
Unfortunately, this vector does not contain expression elements such as a promoter or terminator for proper protein expression. We recommend subcloning your synthetic gene into an expression vector of your choice via restriction enzyme cloning, or we can perform this for you as a custom service.
Yes, you can protect specified regions or restriction enzymes sites that you would not like to be optimized by the the GeneOptimizer process. In the portal, under "Optimize sequence", choose "Protect Restriction Sites" and/or "Protect Sequence Parts" from optimization.
ColE1 is the ori.
Due to our production process, this is not an option we can offer to you. Restriction enzymes are added that generally eliminate most of the standard vectors' multiple cloning site.
Genes are typically cloned into one of our GeneArt pMx standard vectors. These vectors contain
• a multiple cloning site
• an Ori (colE1)
• an antibiotic resistance marker (ampicillin, kanamycin, streptomycin, chloramphenicol); please note standard orders do not guarantee any specific standard vector
There are multiple factors to be considered: CG content, repetition score, direct repeats, and secondary structures, as well the assigned host organism and total length of the gene. All of these things are taken into consideration when deciding whether or not a gene or a part of a sequence is ‘complex'.
The Superspeed delivery option is only available for non-complex genes, up to 3 kb in size. If your gene does not fit these requirements, the option will be greyed out.
Single variants and variant bundles (more than one variant process) are variants of a master gene. The variant criteria are as follows:
Variants may contain up to four of the following modifications:
- 5'/3' deletion of any length with additional 52 nt of new sequence on each side
- 5'/3' modification of a maximum of 52 nt on each side
- 5'/3' extension of a maximum of 52 nt on each side
- Internal modifications of up to 40 nt (a maximum of 3 of these internal modification blocks are allowed)
- Internal deletions of any length
- Modifications (whether internal or 5´/3´) must be separated by at least 100 bp.
If a gene falls outside of these criteria, it must be created as a separate gene synthesis instead of as a variant.
Currently, we do not offer this as a service. Subcloning can be achieved using a restriction enzyme–based vector or via BP/LR clonase reactions into a Gateway vector. Alternatively, the GeneArt Strings DNA Fragments we offer can be directly cloned into a TOPO Zero Blunt vector.
You will receive 5µg lyophilized plasmid (synthesized gene cloned into pMx plasmid), unless otherwise requested. Upon reciept, the DNA can be re-suspended and stored in aliquots at –20 degrees C. You will also receive a CD with sequencing alignment data along with your DNA.
The sizes vary slightly due to the size of the anitibiotic resistance marker. pMA:2.5kb, pMS: 2.6kb, pMC: 2.3kb, pMK: 2.4kb.
The GeneOptimizer process takes a multi-parameter approach to analyze the gene sequence. It will take codon usage, GC content, cryptic splice sites, direct repeats, RNA secondary structures, and instability sequences into account when optimizing a gene sequence.
In theory, no there is no limit. However, very large DNA fragments (greater than 10 kb) need to be cloned into a low-copy vector to ensure genetic stability. Therefore, increasing length leads to increased production time.
We offer a GeneArt Strings DNA Fragments service, which provides custom-made linear, double-stranded DNA fragments up to 1,000 bp in length. Go to www.thermofisher.com, and search for "GeneArt Strings DNA Fragments" to learn more about this product.
Yes, all pMX vectors contain an M13-20 primer binding site and a M13-R primer binding site. The sequences are as follows:
M13-20 forward primer binding site: 5' - TTGTAAAACGACGGCCAG - 3'br/>
M13-R primer binding site: 5' - GGAAACAGCTATGACCATGT - 3'
pMx is the standard GeneArt cloning vector used to clone in your synthetic gene of interest. The “x” stands for the antibiotic of choice. Therefore, pMA stands for ampicillin, pMK stands for kanamycin, pMS stands for spectinomycin, pMC stands for chloramphenicol, and pMZ stands for Zeocin antibiotic. All vectors contain a multiple cloning site, an ORI (colE1), and an antibiotic resistance marker, along with M13-20/M13-R primer binding sites for sequencing.