A Novel and Efficient High-Yield Method for Preparing Bacterial Ghosts
Bacterial ghosts (BGs) are empty cell envelopes possessing native extracellular structures without a cytoplasm and genetic materials. BGs are proposed to have significant prospects in biomedical research as vaccines or delivery carriers. The applications of BGs are often limited by inefficient bacte...
Main Authors: | , , , , , |
---|---|
Format: | Article |
Language: | English |
Published: |
MDPI AG
2021-06-01
|
Series: | Toxins |
Subjects: | |
Online Access: | https://www.mdpi.com/2072-6651/13/6/420 |
id |
doaj-c299a3a7ae114c1cb71d67e6fcb06ad1 |
---|---|
record_format |
Article |
spelling |
doaj-c299a3a7ae114c1cb71d67e6fcb06ad12021-07-01T00:05:25ZengMDPI AGToxins2072-66512021-06-011342042010.3390/toxins13060420A Novel and Efficient High-Yield Method for Preparing Bacterial GhostsYi Ma0Liu Cui1Meng Wang2Qiuli Sun3Kaisheng Liu4Jufang Wang5School of Biology and Biological Engineering, South China University of Technology, Guangzhou 510006, ChinaSchool of Biology and Biological Engineering, South China University of Technology, Guangzhou 510006, ChinaSchool of Biology and Biological Engineering, South China University of Technology, Guangzhou 510006, ChinaSchool of Biology and Biological Engineering, South China University of Technology, Guangzhou 510006, ChinaShenzhen People’s Hospital (The Second Clinical Medical College, Jinan University; The First Affiliated Hospital, Southern University of Science and Technology), Shenzhen 518020, ChinaSchool of Biology and Biological Engineering, South China University of Technology, Guangzhou 510006, ChinaBacterial ghosts (BGs) are empty cell envelopes possessing native extracellular structures without a cytoplasm and genetic materials. BGs are proposed to have significant prospects in biomedical research as vaccines or delivery carriers. The applications of BGs are often limited by inefficient bacterial lysis and a low yield. To solve these problems, we compared the lysis efficiency of the wild-type protein E (E<sub>W</sub>) from phage ΦX174 and the screened mutant protein E (E<sub>M</sub>) in the <i>Escherichia coli</i> BL21(DE3) strain. The results show that the lysis efficiency mediated by protein E<sub>M</sub> was improved. The implementation of the pLysS plasmid allowed nearly 100% lysis efficiency, with a high initial cell density as high as OD<sub>600</sub> = 2.0, which was higher compared to the commonly used BG preparation method. The results of Western blot analysis and immunofluorescence indicate that the expression level of protein E<sub>M</sub> was significantly higher than that of the non-pLysS plasmid. High-quality BGs were observed by SEM and TEM. To verify the applicability of this method in other bacteria, the T7 RNA polymerase expression system was successfully constructed in <i>Salmonella enterica</i> (<i>S. Enterica</i>, SE). A pET vector containing E<sub>M</sub> and pLysS were introduced to obtain high-quality SE ghosts which could provide efficient protection for humans and animals. This paper describes a novel and commonly used method to produce high-quality BGs on a large scale for the first time.https://www.mdpi.com/2072-6651/13/6/420bacterial ghostsΦX174lysis efficiencypLysS |
collection |
DOAJ |
language |
English |
format |
Article |
sources |
DOAJ |
author |
Yi Ma Liu Cui Meng Wang Qiuli Sun Kaisheng Liu Jufang Wang |
spellingShingle |
Yi Ma Liu Cui Meng Wang Qiuli Sun Kaisheng Liu Jufang Wang A Novel and Efficient High-Yield Method for Preparing Bacterial Ghosts Toxins bacterial ghosts ΦX174 lysis efficiency pLysS |
author_facet |
Yi Ma Liu Cui Meng Wang Qiuli Sun Kaisheng Liu Jufang Wang |
author_sort |
Yi Ma |
title |
A Novel and Efficient High-Yield Method for Preparing Bacterial Ghosts |
title_short |
A Novel and Efficient High-Yield Method for Preparing Bacterial Ghosts |
title_full |
A Novel and Efficient High-Yield Method for Preparing Bacterial Ghosts |
title_fullStr |
A Novel and Efficient High-Yield Method for Preparing Bacterial Ghosts |
title_full_unstemmed |
A Novel and Efficient High-Yield Method for Preparing Bacterial Ghosts |
title_sort |
novel and efficient high-yield method for preparing bacterial ghosts |
publisher |
MDPI AG |
series |
Toxins |
issn |
2072-6651 |
publishDate |
2021-06-01 |
description |
Bacterial ghosts (BGs) are empty cell envelopes possessing native extracellular structures without a cytoplasm and genetic materials. BGs are proposed to have significant prospects in biomedical research as vaccines or delivery carriers. The applications of BGs are often limited by inefficient bacterial lysis and a low yield. To solve these problems, we compared the lysis efficiency of the wild-type protein E (E<sub>W</sub>) from phage ΦX174 and the screened mutant protein E (E<sub>M</sub>) in the <i>Escherichia coli</i> BL21(DE3) strain. The results show that the lysis efficiency mediated by protein E<sub>M</sub> was improved. The implementation of the pLysS plasmid allowed nearly 100% lysis efficiency, with a high initial cell density as high as OD<sub>600</sub> = 2.0, which was higher compared to the commonly used BG preparation method. The results of Western blot analysis and immunofluorescence indicate that the expression level of protein E<sub>M</sub> was significantly higher than that of the non-pLysS plasmid. High-quality BGs were observed by SEM and TEM. To verify the applicability of this method in other bacteria, the T7 RNA polymerase expression system was successfully constructed in <i>Salmonella enterica</i> (<i>S. Enterica</i>, SE). A pET vector containing E<sub>M</sub> and pLysS were introduced to obtain high-quality SE ghosts which could provide efficient protection for humans and animals. This paper describes a novel and commonly used method to produce high-quality BGs on a large scale for the first time. |
topic |
bacterial ghosts ΦX174 lysis efficiency pLysS |
url |
https://www.mdpi.com/2072-6651/13/6/420 |
work_keys_str_mv |
AT yima anovelandefficienthighyieldmethodforpreparingbacterialghosts AT liucui anovelandefficienthighyieldmethodforpreparingbacterialghosts AT mengwang anovelandefficienthighyieldmethodforpreparingbacterialghosts AT qiulisun anovelandefficienthighyieldmethodforpreparingbacterialghosts AT kaishengliu anovelandefficienthighyieldmethodforpreparingbacterialghosts AT jufangwang anovelandefficienthighyieldmethodforpreparingbacterialghosts AT yima novelandefficienthighyieldmethodforpreparingbacterialghosts AT liucui novelandefficienthighyieldmethodforpreparingbacterialghosts AT mengwang novelandefficienthighyieldmethodforpreparingbacterialghosts AT qiulisun novelandefficienthighyieldmethodforpreparingbacterialghosts AT kaishengliu novelandefficienthighyieldmethodforpreparingbacterialghosts AT jufangwang novelandefficienthighyieldmethodforpreparingbacterialghosts |
_version_ |
1721349624683298816 |