Basic Study
Copyright ©The Author(s) 2023. Published by Baishideng Publishing Group Inc. All rights reserved.
World J Methodol. Dec 20, 2023; 13(5): 484-491
Published online Dec 20, 2023. doi: 10.5662/wjm.v13.i5.484
Sequential extraction of RNA, DNA and protein from cultured cells of the same group
Ying-Yu Cui
Ying-Yu Cui, Department of Cell Biology, Institute of Medical Genetics, Key Laboratory of Arrhythmias of the Ministry of Education of China, Tongji University School of Medicine, Shanghai 200331, China
Author contributions: Cui YY designed and performed the research, and analyzed the data and wrote the manuscript.
Supported by the Postdoctoral Science Foundation of China, No.2005038300; and the National Natural Science Foundation of China, No. 30671028.
Institutional review board statement: No human and/or animal subjects was involved in the present study.
Institutional animal care and use committee statement: This manuscript does not involve in animal model.
Conflict-of-interest statement: All the authors report no relevant conflicts of interest for this article.
Data sharing statement: No additional data are available.
ARRIVE guidelines statement: This manuscript does not involve in animal model.
Open-Access: This article is an open-access article that was selected by an in-house editor and fully peer-reviewed by external reviewers. It is distributed in accordance with the Creative Commons Attribution NonCommercial (CC BY-NC 4.0) license, which permits others to distribute, remix, adapt, build upon this work non-commercially, and license their derivative works on different terms, provided the original work is properly cited and the use is non-commercial. See: https://creativecommons.org/Licenses/by-nc/4.0/
Corresponding author: Ying-Yu Cui, PhD, Associate Professor, Teacher, Department of Cell Biology, Institute of Medical Genetics, Key Laboratory of Arrhythmias of the Ministry of Education of China, Tongji University School of Medicine, No. 500, Zhennan Road Putuo District, Shanghai 200331, China. yycui@tongji.edu.cn
Received: July 18, 2023
Peer-review started: July 18, 2023
First decision: September 13, 2023
Revised: September 25, 2023
Accepted: October 16, 2023
Article in press: October 16, 2023
Published online: December 20, 2023
Abstract
BACKGROUND

Efficient extraction of nucleic acids and proteins (ENAP) from cells is a prerequisite for precise annotation of gene function, and has become laboratory routine for revealing the mysteries of life. However, cell samples are often from different culture dishes, resulting in inevitable experimental errors and sometimes poor repeatability.

AIM

To explore a method to improve the efficiency of ENAP, minimizing errors in ENAP processes, enhancing the reliability and repeatability of subsequent experimental results.

METHODS

A protocol for the sequential isolation of RNA, DNA, and proteins from the same cultured HepG2 cells using RNAzol reagent is presented here. The first step involves culturing HepG2 cells to the exponential phase, followed by the sequential isolation of RNA, DNA, and proteins from the same cultured cells in the second step. The yield of nucleic acids and proteins is detected in the third step, and their purity and integrity are verified in the last step.

RESULTS

The procedure takes as few as 3-4 d from the start to quality verification and is highly efficient. In contrast to the existing kits and reagents, which are primarily based on independent isolation, this RNAzol reagent-based method is characterized by the sequential isolation of RNA, DNA, and proteins from the same cells, and therefore saves time, and has low cost and high efficiency.

CONCLUSION

The RNA, DNA, and proteins isolated using this method can be used for reverse transcription-polymerase chain reaction, polymerase chain reaction, and western blotting, respectively.

Keywords: Sequential extraction, Ribonucleic acid, Deoxyribonucleic acid, Protein, Cultured cells

Core Tip: Sequential extraction of nucleic acids and proteins from cultured cells of the same group. Life is a way of material (mainly protein and nucleic acid) movement, and health lies in movement. Cell is the most fundamental structural and functional unit of life. Therefore, the effective isolation of nucleic acids and proteins from cells is the foundation and prerequisite for revealing the mysteries of life. However, during laboratory routine for isolation of nucleic acids and proteins, cell samples are often from different culture dishes, usually leading to inevitable experimental errors and sometimes poor repeatability. The present research tries to explore the possibility to simultaneously isolate nucleic acids and proteins from the same sample, while reducing experimental errors and ensuring consistency during experimentation. The present study established a selective protocol for sequential isolation of RNA, DNA and proteins from the same cells with the characteristics of easy operation, rapid extraction and high efficiency. RNAzol reagent was used for the sequential isolation of RNA, DNA, and proteins from the same cultured HepG2 cells, resulting in a novel protocol containing four steps. A protocol for sequential isolation of RNA, DNA and proteins was established and the procedure takes as few as 3-4 d from the start to quality verification and is highly efficient. The quality of RNA, DNA and proteins isolated through sequential isolation protocol can be used for reverse transcription - polymerase chain reaction (PCR), PCR and western blot, respectively. The present procedure is not only easy, rapid and high efficient, but also economical and practical, especially for researchers in developing and underdeveloped countries.