相关实验视频

Updated: Jun 18, 2025

Live Cell Fluorescence Microscopy to Observe Essential Processes During Microbial Cell Growth

Live Cell Fluorescence Microscopy to Observe Essential Processes During Microbial Cell Growth

Published on: November 24, 2017

开发一种基于CRISPR的I-E类型可编程抑制系统,用于微调Bacillus subtilis中的D-Pantothenic酸的代谢流

Chengyao Mao¹, Han Zheng¹, Yifeng Chen¹

¹College of Biotechnology and Bioengineering, Zhejiang University of Technology, Hangzhou, Zhejiang 310014, China.

ACS synthetic biology

|July 31, 2024

概括

在Bacillus subtilis中设计的新CRISPRi系统增强了生物制造的基因调节. 这种改进的CRISPR工具可以提高D-潘托酸的产量,为制造有价值的化学物质提供一种新的方法.

科学领域:

合成生物学合成生物学
微生物工程微生物工程
在CRISPR-Cas系统中.

背景情况:

基于CRISPR的基因调节工具提供了对转录的精确控制,但在细胞毒性和PAM特异性方面存在局限性.
开发新的,毒性较低的CRISPR系统对于扩大其在生物制造和治疗中的应用至关重要.
在细菌细菌中现有的CRISPR工具是有限的,需要新的系统来进行先进的基因工程.

研究的目的:

重建和设计I型CRISPR-Cas系统,用于控制Bacillus subtilis中的基因表达.
通过5'未翻译区域 (UTR) 工程来提高I型CRISPR干扰 (CRISPRi) 系统的有效性.
应用改进的CRISPRi系统,以优化工程B. subtilis.中的D-pantothenic acid (DPA) 生产.

主要方法:

从 Bacillus subtilis 中的 Escherichia coli 中的 I 型 CRISPR-Cas 系统的复制.
对cas基因mRNA的5'未翻译区域 (UTR) 进行工程,以提高CRISPRi系统的效率.
B. subtilis的代谢工程,以加强向β-氨酸和 (R) - 潘托酸的流向,用于DPA生产.
应用CRISPRi系统来控制pdhA表达,以微调代谢流量和TCA循环活动.

主要成果:

在UTR工程之后,I型CRISPRi系统的疗效显著提高.
在B. subtilis的DPA生产中成功进行工程,在摇瓶中达到0.88 g/L,在没有添加前体的料批发发酵中达到12.81 g/L.
通过CRISPRi介导的pdhA表达的控制,证明了对DPA和TCA循环的代谢流量的微调.

结论:

设计的I型CRISPRi系统显著提高了Bacillus subtilis的基因调节能力.
这项研究提供了微调代谢流量的强有力的策略,促进了微生物高效地生产D-潘托酸.
开发的CRISPRi系统和代谢工程方法为在工程生物体中生产化学物质提供了有价值的范式.

关键词:

这种细菌是 Bacillus subtilis.克里斯普里是指克里斯普里.在d-pantothenic 酸中.合成生物学合成生物学类型是IE型.

更多相关视频

In vivo Application of the REMOTE-control System for the Manipulation of Endogenous Gene Expression

In vivo Application of the REMOTE-control System for the Manipulation of Endogenous Gene Expression

Published on: March 29, 2019

In Vitro Selection of Engineered Transcriptional Repressors for Targeted Epigenetic Silencing

In Vitro Selection of Engineered Transcriptional Repressors for Targeted Epigenetic Silencing

Published on: May 5, 2023

相关概念视频

Prokaryotic Transcriptional Activators and Repressors

Prokaryotic Transcriptional Activators and Repressors

The organization of prokaryotic genes in their genome is notably different from that of eukaryotes. Prokaryotic genes are organized, such that the genes for proteins involved in the same biochemical process or function are located together in groups. This group of genes, along with their regulatory elements, are collectively known as an operon. The functional genes in an operon are transcribed together to give a single strand of mRNA known as polycistronic mRNA.
Transcription of prokaryotic...

Riboswitches

Riboswitches

Riboswitches are non-coding mRNA domains that regulate the transcription and translation of downstream genes without the help of proteins. Riboswitches bind directly to a metabolite and can form unique stem-loop or hairpin structures in response to the amount of the metabolite present. They have two distinct regions – a metabolite-binding aptamer and an expression platform.
The aptamer has high specificity for a particular metabolite which allows riboswitches to specifically regulate...

CRISPR and crRNAs

CRISPR and crRNAs

Bacteria and archaea are susceptible to viral infections just like eukaryotes; therefore, they have developed a unique adaptive immune system to protect themselves. Clustered regularly interspaced short palindromic repeats and CRISPR-associated proteins (CRISPR-Cas) are present in more than 45% of known bacteria and 90% of known archaea.
The CRISPR-Cas system stores a copy of foreign DNA in the host genome and uses it to identify the foreign DNA upon reinfection. CRISPR-Cas has three different...