作者: 有發(fā)明天才 時(shí)間: 2025-3-21 23:16
Use of a Golden Gate Plasmid Set Enabling Scarless MoClo-Compatible Transcription Unit Assembly,erate transcription units. Typically, type IIS enzymes are used, which generate four nucleotide overhangs. This results in small scar sequences in hierarchical DNA assemblies, which can affect the functionality of transcription units. However, there are enzymes that generate three nucleotide overhan作者: 半球 時(shí)間: 2025-3-22 01:34 作者: inveigh 時(shí)間: 2025-3-22 07:54
Validation of Golden Gate Assemblies Using Highly Multiplexed Nanopore Amplicon Sequencing,libraries, resulting in assemblies of high combinatorial complexity where sequencing again becomes mandatory. Here, we present a detailed protocol for the use of a highly multiplexed dual barcode amplicon sequencing using the Nanopore sequencing platform for in-house sequence validation. The workflo作者: Ptosis 時(shí)間: 2025-3-22 09:47
Modular Golden Gate Assembly of Linear DNA Templates for Cell-Free Prototyping,tion of a repressor–promoter pair. By eliminating time-consuming transformation and cloning steps in cells and by taking advantage of modular cloning toolboxes, our cell-free prototyping workflow can produce data for large numbers of new assembled constructs within a single day.作者: convert 時(shí)間: 2025-3-22 14:29 作者: convert 時(shí)間: 2025-3-22 18:07 作者: BIDE 時(shí)間: 2025-3-22 23:33 作者: Lumbar-Spine 時(shí)間: 2025-3-23 03:37
Rosemarie Klemm,Dietrich D. Klemming the type of part that needs to be cloned, (2) designing primers for amplification, (3) performing polymerase chain reaction (PCR) amplification, (4) cloning of the fragments using Golden Gate cloning, and finally (5) sequencing of the part. For large standard parts, it is preferable to first clo作者: coagulation 時(shí)間: 2025-3-23 07:12 作者: eustachian-tube 時(shí)間: 2025-3-23 12:49
Rollenkonzepte dynamischer Organisationen,, and modular cloning indexed plasmid library assembly. These upgrades enable a broad pool of users with varying levels of experience to readily implement advanced Golden Gate applications using low-cost, open-source lab robotics.作者: Tremor 時(shí)間: 2025-3-23 16:13 作者: 思考 時(shí)間: 2025-3-23 21:05
Formulierung der Forschungsfragen,tion of a repressor–promoter pair. By eliminating time-consuming transformation and cloning steps in cells and by taking advantage of modular cloning toolboxes, our cell-free prototyping workflow can produce data for large numbers of new assembled constructs within a single day.作者: Arb853 時(shí)間: 2025-3-23 22:59
,Zur Struktur der Besch?ftigten,nhance solubility and expression. The system allows visual candidate screening through mScarlet fluorescence and solubility tags are removable via TEV protease cleavage. We provide a comprehensive protocol encompassing oligonucleotide design, cloning, expression, His-tag affinity chromatography, and作者: 復(fù)習(xí) 時(shí)間: 2025-3-24 05:27
https://doi.org/10.1007/978-3-7091-1232-8ly de-modify methylated or hydroxymethylated cytosines on phage DNA. The temporal removal of cytosine modification ultimately enables efficient DNA cleavage by Cas enzymes and facilitates mutagenesis. To streamline the application of the coupled TET-CRISPR-Cas system, we use Golden Gate cloning for 作者: 清唱?jiǎng)?nbsp; 時(shí)間: 2025-3-24 09:47
https://doi.org/10.1007/978-90-481-3040-5ed plasmid backbone, for rapid and simple production of complex, multi-part assemblies. Insertion of a gene for superfolder green fluorescent protein, with selection by tetracycline resistance, into . strain MG1655 is used as an example but in principle the method can be tailored for virtually any m作者: 聲音刺耳 時(shí)間: 2025-3-24 11:19 作者: Irritate 時(shí)間: 2025-3-24 15:15 作者: radiograph 時(shí)間: 2025-3-24 22:16
Selection of Fusion-Site Overhang Sets for High-Fidelity and High-Complexity Golden Gate Assembly,ired order. Traditionally, fusion-site sequences are selected by using validated sets of overhang sequences or by applying a handful of semi-empirical rules to guide overhang choice. While these approaches allow dependable assembly of 6–8 fragments in one pot, recent work has demonstrated that compr作者: 新陳代謝 時(shí)間: 2025-3-25 00:09 作者: Flirtatious 時(shí)間: 2025-3-25 03:47
Using ApE for In Silico Golden Gate Cloning, of the restriction fragment overhangs to minimize undesired products and to generate the desired junctions. The ApE (A plasmid Editor) software package can assist in silico design of input fragments or to generate expected assembly products.作者: Carminative 時(shí)間: 2025-3-25 09:03
Standardized Golden Gate Assembly Metadata Representation Using SBOL, to engineer biological systems is by modifying their DNA. A common workflow involves creating new DNA parts through synthesis and then using them in combination with other parts through assembly. Assembly standards such as MoClo, Phytobricks, and Loop are based on Golden Gate, and provide a framewo作者: Crepitus 時(shí)間: 2025-3-25 13:11
Use of a Golden Gate Plasmid Set Enabling Scarless MoClo-Compatible Transcription Unit Assembly,d construction of transcription units and multi-gene constructs. Importantly, its modular structure makes it compatible with laboratory automation, allowing for systematic and highly complex DNA assembly. Golden Gate cloning relies on type IIS enzymes that cleave an adjacent undefined sequence motif作者: mercenary 時(shí)間: 2025-3-25 19:29
Biofoundry-Assisted Golden Gate Cloning with AssemblyTron,typing. However, compared to robotic implementation, manual Golden Gate implementation is cumbersome, error-prone, and inconsistent for complex assembly designs. AssemblyTron is an open-source python package that provides an affordable automation solution using open-source OpenTrons OT-2 lab robots.作者: Morbid 時(shí)間: 2025-3-25 20:27 作者: 可以任性 時(shí)間: 2025-3-26 01:28
Validation of Golden Gate Assemblies Using Highly Multiplexed Nanopore Amplicon Sequencing,gher order assemblies allows engineering principles to be applied to biological systems. The basic parts, typically stored in Level 0 plasmids, are sequence validated by the method of choice and can be combined into higher order assemblies on demand. Higher order assemblies are typically transcripti作者: Fortify 時(shí)間: 2025-3-26 05:54 作者: Eviction 時(shí)間: 2025-3-26 10:42
Construction of Small Genes with Repetitive Elements Using Oligo Extension and Golden Gate Assemblyr delays by DNA synthesis vendors. Here, we describe a method for the assembly of small synthetic genes with repetitive elements: First, a gene of interest is split in silico into small synthons of up to 80 base pairs flanked by Golden Gate-compatible overhangs. Then synthons are made by oligo exten作者: travail 時(shí)間: 2025-3-26 14:31
Simplifying Recombinant Protein Production: Combining Golden Gate Cloning with a Standardized Protend structural analyses of the purified samples. The demand for substantial biomolecule quantities often presents challenges, particularly for eukaryotic proteins. . expression systems have evolved to address these issues, offering advanced features such as solubility tags, posttranslational modifica作者: BAIL 時(shí)間: 2025-3-26 17:59 作者: 角斗士 時(shí)間: 2025-3-27 00:16 作者: 畫布 時(shí)間: 2025-3-27 02:14 作者: 保存 時(shí)間: 2025-3-27 06:57 作者: 男生如果明白 時(shí)間: 2025-3-27 12:56
Utilizing Golden Gate Assembly to Streamline CRISPR-Cas/NgTET-Based Phage Mutagenesis,age mutagenesis tools are required to customize the phages for a specific application and generate, in addition to that, so-called designer phages. CRISPR-Cas technique is used in various organisms to perform targeted mutagenesis. Yet, its efficacy is notably limited for phage mutagenesis due to the作者: 帶來的感覺 時(shí)間: 2025-3-27 17:07 作者: linguistics 時(shí)間: 2025-3-27 19:56
Rosemarie Klemm,Dietrich D. Klemmning for DNA assembly, are designed to facilitate assembly of multigene constructs from libraries of standard parts through a series of streamlined one-pot assembly reactions. Standard parts consist of the DNA sequence of a genetic element of interest such as a promoter, coding sequence, or terminat作者: 指耕作 時(shí)間: 2025-3-28 01:31
https://doi.org/10.1007/978-3-663-05453-5hetic biology. These systems are simple to use, efficient, and allow users to assemble multigene constructs by performing a series of one-pot assembly steps, starting from libraries of cloned and sequenced parts. The efficiency of these systems also facilitates the generation of libraries of constru作者: 催眠藥 時(shí)間: 2025-3-28 02:22
Fragestellung, Konzept und Aufbau der Studieired order. Traditionally, fusion-site sequences are selected by using validated sets of overhang sequences or by applying a handful of semi-empirical rules to guide overhang choice. While these approaches allow dependable assembly of 6–8 fragments in one pot, recent work has demonstrated that compr作者: Perceive 時(shí)間: 2025-3-28 07:51
Zusammensetzung des Steinkohlenteers, makes them particularly amenable to high-throughput automation, facilitating the generation of thousands of constructs in a massively parallel manner. One potential bottleneck in this process is the design of these constructs. There are multiple parameters that must be considered during the design 作者: 北京人起源 時(shí)間: 2025-3-28 12:22
Evolution of Rapidly Rotating B-Type Stars of the restriction fragment overhangs to minimize undesired products and to generate the desired junctions. The ApE (A plasmid Editor) software package can assist in silico design of input fragments or to generate expected assembly products.作者: separate 時(shí)間: 2025-3-28 15:04 作者: figure 時(shí)間: 2025-3-28 21:03
,Was Sie aus diesem Buch lernen k?nnen,d construction of transcription units and multi-gene constructs. Importantly, its modular structure makes it compatible with laboratory automation, allowing for systematic and highly complex DNA assembly. Golden Gate cloning relies on type IIS enzymes that cleave an adjacent undefined sequence motif作者: 載貨清單 時(shí)間: 2025-3-29 02:59 作者: Outspoken 時(shí)間: 2025-3-29 06:01 作者: Malaise 時(shí)間: 2025-3-29 08:55 作者: META 時(shí)間: 2025-3-29 11:29 作者: 擁護(hù)者 時(shí)間: 2025-3-29 18:02
Methodologische Vorbemerkungen,r delays by DNA synthesis vendors. Here, we describe a method for the assembly of small synthetic genes with repetitive elements: First, a gene of interest is split in silico into small synthons of up to 80 base pairs flanked by Golden Gate-compatible overhangs. Then synthons are made by oligo exten作者: 假裝是你 時(shí)間: 2025-3-29 22:51
,Zur Struktur der Besch?ftigten,nd structural analyses of the purified samples. The demand for substantial biomolecule quantities often presents challenges, particularly for eukaryotic proteins. . expression systems have evolved to address these issues, offering advanced features such as solubility tags, posttranslational modifica作者: 地名詞典 時(shí)間: 2025-3-29 23:58 作者: 滲透 時(shí)間: 2025-3-30 04:47
https://doi.org/10.1007/978-3-663-05454-2 cloning. While currently offering the most affordable source of synthetic DNA, oligo pools also come with limitations such as a maximum synthesis length (approximately 350 bases), a higher error rate compared to alternative synthesis methods, and the presence of truncated molecules in the pool due 作者: DIS 時(shí)間: 2025-3-30 10:20
Ontwikkeling en veroudering van de stem,r sequences that can be engineered for custom target sites. These arrays are transcribed into precursor CRISPR RNAs (pre-crRNAs) that undergo maturation steps to form individual CRISPR RNAs (crRNAs). Each crRNA contains a single spacer that identifies the target cleavage site for a large variety of 作者: 注意 時(shí)間: 2025-3-30 14:38 作者: GROG 時(shí)間: 2025-3-30 20:22
https://doi.org/10.1007/978-3-7091-1232-8age mutagenesis tools are required to customize the phages for a specific application and generate, in addition to that, so-called designer phages. CRISPR-Cas technique is used in various organisms to perform targeted mutagenesis. Yet, its efficacy is notably limited for phage mutagenesis due to the作者: defray 時(shí)間: 2025-3-31 00:06
https://doi.org/10.1007/978-90-481-3040-5integration of a fragment of DNA by homologous recombination (known as recombineering). In contrast to the traditional recombineering method, the integrated fragment for Gene Doctoring is supplied on a donor plasmid rather than as a linear DNA. This protects the DNA from degradation, facilitates tra作者: Paradox 時(shí)間: 2025-3-31 01:26
Definition of Anatomical Features,esent the G-GArden tool, which assists with the design of oligodeoxynucleotides and overhangs for scarless assembly strategies. We propose that the presented procedures are suitable for many applications in different bacteria, which are related to . and beyond.作者: 吸氣 時(shí)間: 2025-3-31 08:20 作者: 一再煩擾 時(shí)間: 2025-3-31 11:41 作者: assent 時(shí)間: 2025-3-31 16:58 作者: angiography 時(shí)間: 2025-3-31 17:33
https://doi.org/10.1007/978-3-322-89681-0ield of synthetic biology. The use of this technology allows miniaturization and parallelization of molecular reactions in a tip-free manner, making it sustainable by reducing plastic waste and reagent usage. Here, we provide a step-by-step protocol for performing and parallelizing Golden Gate cloning reactions in 1?μL total volume.作者: AVOW 時(shí)間: 2025-3-31 23:43 作者: ACRID 時(shí)間: 2025-4-1 02:42 作者: Working-Memory 時(shí)間: 2025-4-1 09:03
Golden Gate Cloning of Multigene Constructs Using the Modular Cloning System MoClo, The assembly strategy is then defined following a set of standard rules. Multigene constructs are then assembled using a series of one-pot assembly steps with the set of identified parts and vectors.作者: Bravado 時(shí)間: 2025-4-1 10:32 作者: Misnomer 時(shí)間: 2025-4-1 14:31
Automation and Miniaturization of Golden Gate DNA Assembly Reactions Using Acoustic Dispensers,ield of synthetic biology. The use of this technology allows miniaturization and parallelization of molecular reactions in a tip-free manner, making it sustainable by reducing plastic waste and reagent usage. Here, we provide a step-by-step protocol for performing and parallelizing Golden Gate cloning reactions in 1?μL total volume.作者: 銀版照相 時(shí)間: 2025-4-1 19:07
Combining Oligo Pools and Golden Gate Cloning to Create Protein Variant Libraries or Guide RNA Librwork within the Yeast Toolkit Golden Gate scheme, but are in principle compatible with any other Golden Gate-based modular cloning toolkit and extendable to other restriction enzyme-based cloning methods beyond Golden Gate. Our methods yield high-quality, affordable, in-house variant libraries.
