pNY326短短芽胞杆菌表达载体Brevibacillus expression plasmid BioVector NTCC中国质粒载体菌种细胞基因保藏中心

pNY326短芽胞杆菌表达载体Brevibacillus expression vector

Cat#: BioVector-HB111                    

Description

TheBrevibacillus (Bacillus brevis) Expression System is an efficient system for producing high

yields of secreted protein. Using this system, it is possible to produce an unprecedented

number of heterologous proteins. TheBrevibacillus Expression System offers the following

advantages, and is particularly effective in the production of secretory proteins.

・Extracellular secretion of large amounts of protein

・Almost no detectable protease activity

・Produces active proteins

・Simple procedures for genetic manipulation and culturing

・Safe host bacteria

Examples of protein production using this system are shown in Table 1. High expression levels

were obtained for a variety of proteins (enzymes, antigens, and cytokines), regardless of gene

origin (bacteria, archaea and eukaryotes). In particular, secretory proteins of eukaryotic origin

typically contain S-S bonds for biological activity, and it is generally difficult to produce these

proteins using other prokaryotic expression systems. The characteristic secretory production

ofBrevibacillus allows high-efficiency production, even for proteins with S-S bonds.

It is possible to construct the expression vector using a shuttle vector inE. coli or by directly

introducing the DNA ligation solution into the expression host. Target protein may be obtained

by culturing transformed host bacteria in test tubes or flasks (two types of media can be

used; see Section VI. 9 for medium composition), and collecting protein from the culture

supernatant. Homogenization of bacteria is unnecessary; the target protein is obtained from

the clear supernatant fraction following centrifugation (to pellet the bacterial cells) and may

be purified using standard techniques.

Components

Brevibacillus Expression System II (Cat. #HB200)

● Expression Vectors

pNY326 DNA  10 μg (Cat. #HB111)

pNCMO2 DNA  10 μg (Cat. #HB112)

● Control Vector

pNY326-BLA DNA  1 μg (Cat. #HB114)

● Competent Cells

Brevibacillus Competent Cells (Cat. #HB116)

・Brevibacillus Competent Cells  100 μl x 10

・MT medium  1 ml x 10

・Solution A  1 ml

・Solution B  1 ml x 2

Brevibacillus (Bacillus brevis ) Expression System是能够高效表达分泌型和优良蛋白质的表达系统，并能生产大量的异源蛋白质。本系统具有如下优点，尤其适用于生产分泌型蛋白质。 pNY326能够在B. choshinensis 细胞中复制，它的大小(3.4 kb)比pNCMO2(5.2 kb)小得多，且启动子活性也比pNCMO2弱。这样，即便是表达对宿主细胞有毒性的蛋白质，pNY326也能保持稳定，在某些情况下，也由于转化株的稳定生长可以获得高产量的蛋白质。稳定的产量使得pNY326尤其适合大规模培养。使用B.choshinensis 宿主可一步构建表达质粒，要保证足够高的转化效率，建议使用Brevibacillus Competent Cells·生产大量的胞外分泌型蛋白质；·无蛋白酶活性；·生产有活性的蛋白质；·易于培养和灭菌； ·简单的基因技术； ·安全的宿主菌。使用本系统，不论是哪种基因起源(细菌，古细菌，真核生物)，各种蛋白质(酶，抗原和细胞因子)都获得了高水平的表达。尤其是真核起源的分泌型蛋白质一般具有S-S键结构，本系统也能进行具有活性的表达，这是使用其他原核表达系统都很难做到的。构建表达载体时，可以是E.coli 的穿梭载体，也可以是直接将DNA连接液转化到表达宿主中。然后目的蛋白质可通过转化后宿主菌的培养，收集培养上清液获得。不需要细菌匀浆破碎的过程，离心(去除菌细胞)后取上清，然后使用常规技术进行蛋白质的纯化。

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