电穿孔和裂解海洋微藻凯伦藻用于 RNA 提取和扩增。
Electroporation and lysis of marine microalga Karenia brevis for RNA extraction and amplification.
机构信息
Sensors Development Group, National Oceanography Centre, Southampton, UK.
出版信息
J R Soc Interface. 2011 Apr 6;8(57):601-8. doi: 10.1098/rsif.2010.0445. Epub 2010 Nov 17.
Abstract
We describe here a simple device for dielectrophoretic concentration of marine microalga Karenia brevis non-motile cells, followed by electric field-mediated lysis for RNA extraction. The lysate was purified using magnetic beads and pure RNA extracted. RNA quality was assessed off-chip by nucleic acid sequence-based amplification and the optimum conditions for lysis were determined. This procedure will form part of an integrated microfluidic system that is being developed with sub-systems for performing cell concentration and lysis, RNA extraction/purification and real-time quantitative RNA detection. The integrated system and its components could be used for a large range of applications including in situ harmful algal bloom detection, transcriptomics and point-of-care diagnostics.
摘要
我们在这里描述了一种简单的设备，用于进行电活性泳富集海洋微藻无毒细胞，然后通过电场介导的裂解进行 RNA 提取。裂解物用磁珠进行纯化，并提取纯 RNA。通过基于核酸序列的扩增对 RNA 质量进行了芯片外评估，并确定了最佳的裂解条件。该程序将成为正在开发的集成微流控系统的一部分，该系统具有用于进行细胞浓缩和裂解、RNA 提取/纯化以及实时定量 RNA 检测的子系统。该集成系统及其组件可用于多种应用，包括原位有害藻华检测、转录组学和即时诊断。
相似文献
1
Electroporation and lysis of marine microalga Karenia brevis for RNA extraction and amplification.电穿孔和裂解海洋微藻凯伦藻用于 RNA 提取和扩增。
J R Soc Interface. 2011 Apr 6;8(57):601-8. doi: 10.1098/rsif.2010.0445. Epub 2010 Nov 17.
2
Real-time isothermal RNA amplification of toxic marine microalgae using preserved reagents on an integrated microfluidic platform.利用集成微流控平台上保存的试剂对有毒海洋微藻进行实时等温 RNA 扩增。
Analyst. 2013 Jan 21;138(2):593-602. doi: 10.1039/c2an36464f.
3
Detection and quantification of the toxic microalgae Karenia brevis using lab on a chip mRNA sequence-based amplification.使用基于芯片实验室mRNA序列扩增技术检测和定量有毒微藻短裸甲藻。
J Microbiol Methods. 2017 Aug;139:189-195. doi: 10.1016/j.mimet.2017.06.008. Epub 2017 Jun 8.
4
Integrated sample-to-detection chip for nucleic acid test assays.用于核酸检测分析的集成式样本到检测芯片。
Biomed Microdevices. 2016 Jun;18(3):44. doi: 10.1007/s10544-016-0069-8.
5
Detecting Harmful Algal Blooms with Isothermal Molecular Strategies.利用等温分子策略探测有害藻华。
Trends Biotechnol. 2019 Dec;37(12):1278-1281. doi: 10.1016/j.tibtech.2019.07.003. Epub 2019 Aug 6.
6
Digital Microfluidic Approach for Efficient Electroporation with High Productivity: Transgene Expression of Microalgae without Cell Wall Removal.用于高效电穿孔且具有高生产率的数字微流控方法：无需去除细胞壁的微藻转基因表达。
Anal Chem. 2015 Jul 7;87(13):6592-9. doi: 10.1021/acs.analchem.5b00725. Epub 2015 Jun 11.
7
Pathogenic Bacteria Detection Using RNA-Based Loop-Mediated Isothermal-Amplification-Assisted Nucleic Acid Amplification via Droplet Microfluidics.基于 RNA 的环介导等温扩增辅助的液滴微流控技术用于致病性细菌检测。
ACS Sens. 2019 Apr 26;4(4):841-848. doi: 10.1021/acssensors.8b01206. Epub 2019 Mar 25.
8
A novel portable filtration system for sampling and concentration of microorganisms: Demonstration on marine microalgae with subsequent quantification using IC-NASBA.一种新型便携式微生物采样浓缩过滤系统：海洋微藻的应用实例及后续采用 IC-NASBA 定量分析。
Harmful Algae. 2018 May;75:94-104. doi: 10.1016/j.hal.2018.03.006. Epub 2018 Apr 25.
9
Establishment and application of hyperbranched rolling circle amplification coupled with lateral flow dipstick for the sensitive detection of Karenia mikimotoi.建立和应用超分支滚环扩增结合侧流纸条用于敏感检测米氏凯伦藻。
Harmful Algae. 2019 Apr;84:151-160. doi: 10.1016/j.hal.2019.03.015. Epub 2019 Apr 10.
10
A microfluidic liquid phase nucleic acid purification chip to selectively isolate DNA or RNA from low copy/single bacterial cells in minute sample volume followed by direct on-chip quantitative PCR assay.一种微流控液相核酸纯化芯片，可从低拷贝/单个细菌细胞中选择性分离 DNA 或 RNA，样品体积极小，随后可直接在芯片上进行定量 PCR 分析。
Anal Chem. 2013 Feb 5;85(3):1484-91. doi: 10.1021/ac3026509. Epub 2013 Jan 17.
引用本文的文献
1
The Pre-Polarization and Concentration of Cells near Micro-Electrodes Using AC Electric Fields Enhances the Electrical Cell Lysis in a Sessile Drop.利用交流电场对微电极附近细胞进行预极化和浓缩可增强静态液滴中的电穿孔细胞裂解。
Biosensors (Basel). 2025 Jan 6;15(1):22. doi: 10.3390/bios15010022.
2
A low-voltage alternant direct current electroporation chip for ultrafast releasing the genome DNA of Helicobacter pylori bacterium.一种用于快速释放幽门螺旋杆菌基因组 DNA 的低压交变直流电穿孔芯片。
Mikrochim Acta. 2024 Jan 30;191(2):116. doi: 10.1007/s00604-024-06187-6.
3
Separation, Characterization, and Handling of Microalgae by Dielectrophoresis.基于介电电泳的微藻分离、表征及处理
Microorganisms. 2020 Apr 9;8(4):540. doi: 10.3390/microorganisms8040540.
4
Electrochemical cell lysis of gram-positive and gram-negative bacteria: DNA extraction from environmental water samples.革兰氏阳性菌和革兰氏阴性菌的电化学细胞裂解：从环境水样中提取DNA
Electrochim Acta. 2020 Apr 1;338:135864. doi: 10.1016/j.electacta.2020.135864.
5
Molecular harvesting with electroporation for tissue profiling.电穿孔法进行分子提取以进行组织剖析。
Sci Rep. 2019 Oct 31;9(1):15750. doi: 10.1038/s41598-019-51634-7.
6
Microfluidic techniques for enhancing biofuel and biorefinery industry based on microalgae.基于微藻的用于提升生物燃料和生物精炼产业的微流控技术。
Biotechnol Biofuels. 2019 Feb 15;12:33. doi: 10.1186/s13068-019-1369-z. eCollection 2019.
7
Microfluidic Sample Preparation for Single Cell Analysis.用于单细胞分析的微流控样品制备
Anal Chem. 2016 Jan 5;88(1):354-80. doi: 10.1021/acs.analchem.5b04077. Epub 2015 Dec 3.
8
Microfluidic electroporation for cellular analysis and delivery.微流控电穿孔用于细胞分析和药物递送。
Lab Chip. 2013 Oct 7;13(19):3803-21. doi: 10.1039/c3lc50566a.
9
Flow process for electroextraction of total proteins from microalgae.从微藻中电萃取总蛋白的流程。
J Membr Biol. 2013 Oct;246(10):751-60. doi: 10.1007/s00232-013-9542-y. Epub 2013 Apr 11.
本文引用的文献
1
Nucleic Acid-based Detection of Bacterial Pathogens Using Integrated Microfluidic Platform Systems.基于核酸的集成微流控平台系统检测细菌病原体。
Sensors (Basel). 2009;9(5):3713-44. doi: 10.3390/s90503713. Epub 2009 May 18.
2
Marine chemical technology and sensors for marine waters: potentials and limits.海洋化学技术与海洋水质传感器：潜力与限制。
Ann Rev Mar Sci. 2009;1:91-115. doi: 10.1146/annurev.marine.010908.163817.
3
Electrochemical cell lysis device for DNA extraction.电化学细胞裂解装置，用于 DNA 提取。
Lab Chip. 2010 Mar 7;10(5):626-33. doi: 10.1039/b916606h. Epub 2009 Dec 17.
4
Microfluidic sample preparation: cell lysis and nucleic acid purification.微流控样品制备：细胞裂解和核酸纯化。
Integr Biol (Camb). 2009 Oct;1(10):574-86. doi: 10.1039/b905844c. Epub 2009 Aug 25.
5
Nucleic acid extraction techniques and application to the microchip.核酸提取技术及其在微芯片上的应用。
Lab Chip. 2009 Sep 7;9(17):2484-94. doi: 10.1039/b907652m. Epub 2009 Jun 22.
6
Cellular membrane potentials induced by alternating fields.交变电场诱导的细胞膜电位。
Biophys J. 1992 Dec;63(6):1632-42. doi: 10.1016/S0006-3495(92)81740-X.
7
Perspective on optical biosensors and integrated sensor systems.光学生物传感器与集成传感器系统的展望。
Anal Chem. 2009 Jan 15;81(2):519-26. doi: 10.1021/ac8016289.
8
Microfluidic device for dielectrophoresis manipulation and electrodisruption of respiratory pathogen Bordetella pertussis.用于对呼吸道病原体百日咳博德特氏菌进行介电电泳操控和电破碎的微流控装置。
IEEE Trans Biomed Eng. 2008 Oct;55(10):2426-32. doi: 10.1109/TBME.2008.923148.
9
Purification of nucleic acids in microfluidic devices.微流控设备中核酸的纯化
Anal Chem. 2008 Sep 1;80(17):6472-9. doi: 10.1021/ac8014998.
10
Microfluidic single-cell analysis of intracellular compounds.细胞内化合物的微流控单细胞分析
J R Soc Interface. 2008 Oct 6;5 Suppl 2(Suppl 2):S139-50. doi: 10.1098/rsif.2008.0233.focus.
Zotero 插件