DNA shuffling is a powerful tool to develop libraries of variants for protein engineering. Here, we present a protocol to use our freely available and easy-to-use computer program, Shuffle Optimizer. Shuffle Optimizer is written in the Python computer language and increases the nucleotide homology between two pieces of DNA desired to be shuffled together without changing the amino acid sequence. In addition we also include sections on optimal primer design for DNA shuffling and library construction, a small-volume ultrasonicator method to create sheared DNA, and finally a method to reassemble the sheared fragments and recover and clone the library. The Shuffle Optimizer program and these protocols will be useful to anyone desiring to perform any of the nucleotide homology-dependent shuffling methods.
Download full-text PDF |
Source |
|---|---|
| http://dx.doi.org/10.1007/978-1-4939-6343-0_3 | DOI Listing |
Publication Analysis
Top Keywords
Similar Publications
Research on machine vision online monitoring system for egg production and quality in cage environment.
Poult Sci
November 2024
College of Engineering, South China Agricultural University, Guangzhou, China; State Key Laboratory of Livestock and Poultry Breeding, Guangzhou, China. Electronic address:
In the domain of egg production, the application of automation technologies is essential for boosting productivity and quality. This study introduces an online monitoring system designed for egg quality assessment within caged environments, incorporating a robotic patrol system for egg localization and a fixed video stream for quality analysis. The project involved upgrading traditional henhouses with enhanced wireless connectivity and developing data transmission techniques for video streams and image data.
View Article and Find Full Text PDFResearch on Fine-Tuning Optimization Strategies for Large Language Models in Tabular Data Processing.
Biomimetics (Basel)
November 2024
School of Management Science and Engineering, Beijing Information Science and Technology University, Beijing 100192, China.
Recent advancements in natural language processing (NLP) have been significantly driven by the development of large language models (LLMs). Despite their impressive performance across various language tasks, these models still encounter challenges when processing tabular data. This study investigates the optimization of fine-tuning strategies for LLMs specifically in the context of tabular data processing.
View Article and Find Full Text PDFPapain expression in the cytoplasm by T7-promoter engineering and co-expression with human protein disulfide isomerase (PDI) and thiol peroxidase (GPx7) genes.
Appl Environ Microbiol
November 2024
Center for Biotechnology and Interdisciplinary Studies, Rensselaer Polytechnic Institute, Troy, New York, USA.
Unlabelled: Difficulties exist in obtaining full-length, correctly folded, and soluble papain or papain-like proteases that necessitate the exploration of alternative strategies. This study describes the development of an strain capable of producing soluble papain without the need for complex and time-consuming refolding steps. To enhance the production of soluble papain, engineered T7 promoters and a recombinant papain translationally fused with varying tags were constructed.
View Article and Find Full Text PDFIncreased cytoplasmic expression of PETase enzymes in E. coli.
Microb Cell Fact
November 2024
Department of Chemical, Paper and Biomedical Engineering, Miami University, 650 E. High St., Engineering Building 64, Oxford, OH, 45056, USA.
Background: Depolymerizing polyethylene terephthalate (PET) plastics using enzymes, such as PETase, offers a sustainable chemical recycling route. To enhance degradation, many groups have sought to engineer PETase for faster catalysis on PET and elevated stability. Considerably less effort has been focused toward expressing large quantities of the enzyme, which is necessary for large-scale application and widespread use.
View Article and Find Full Text PDFOptimized production of a truncated form of the recombinant neuraminidase of influenza virus in Escherichia coli as host with suitable functional activity.
Microb Cell Fact
November 2024
Virology Department, Pasteur Institute of Iran, Tehran, Iran.
Background: To discover effective drugs for treating Influenza (a disease with high annual mortality), large amounts of recombinant neuraminidase (NA) with suitable catalytic activity are needed. However, the functional activity of the full-length form of this enzyme in the bacterial host (as producing cells with a low cost) in a soluble form is limited. Thus, in the present study, a truncated form of the neuraminidase (derived from California H1N1 influenza strain) was designed, then biosynthesized in Escherichia coli BL21 (DE3), Shuffle T7, and SILEX systems.
View Article and Find Full Text PDFWant AI Summaries of new PubMed Abstracts delivered to your In-box?
Enter search terms and have AI summaries delivered each week - change queries or unsubscribe any time!