An Efficient and Universal Protoplast Isolation Protocol Suitable for Transient Gene Expression Analysis and Single-Cell RNA Sequencing

doi:10.3390/ijms23073419

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	An Efficient and Universal Protoplast Isolation Protocol Suitable for Transient Gene Expression Analysis and Single-Cell RNA Sequencing
	Wang, Juanjuan 1; Wang, Yang 1; Lu, Tianfeng ; Yang, Xia ; Liu, Jing ; Dong, Yang 1; Wang, Yinzheng 1
刊名	INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES
	2022
卷号	23 期号:7
关键词	protoplast isolation cell walls Chirita pumila protocol transient gene expression single-cell RNA sequencing
DOI	10.3390/ijms23073419
文献子类	Article
英文摘要	The recent advent of single-cell RNA sequencing (scRNA-seq) has enabled access to the developmental landscape of a complex organ by monitoring the differentiation trajectory of every specialized cell type at the single-cell level. A main challenge in this endeavor is dissociating plant cells from the rigid cell walls and some species are recalcitrant to such cellular isolation. Here, we describe the establishment of a simple and efficient protocol for protoplast preparation in Chirita pumila, which includes two consecutive digestion processes with different enzymatic buffers. Using this protocol, we generated viable cell suspensions suitable for an array of expression analyses, including scRNA-seq. The universal application of this protocol was further tested by successfully isolating high-quality protoplasts from multiple organs (petals, fruits, tuberous roots, and gynophores) from representative species on the key branches of the angiosperm lineage. This work provides a robust method in plant science, overcoming barriers to isolating protoplasts in diverse plant species and opens a new avenue to study cell type specification, tissue function, and organ diversification in plants.
学科主题	Biochemistry & Molecular Biology ; Chemistry, Multidisciplinary
电子版国际标准刊号	1422-0067
出版地	BASEL
WOS关键词	MESOPHYLL PROTOPLASTS ; ARABIDOPSIS ; PECTIN ; MORPHOGENESIS ; EVOLUTION ; SYSTEM
WOS研究方向	Science Citation Index Expanded (SCI-EXPANDED)
语种	英语
出版者	MDPI
WOS记录号	WOS:000781113700001
资助机构	Natural Science Foundation of China [32170227, 31970239]
内容类型	期刊论文
源URL	[http://ir.ibcas.ac.cn/handle/2S10CLM1/28746]
专题	系统与进化植物学国家重点实验室
作者单位	1.Chinese Acad Sci, Inst Bot, State Key Lab Systemat & Evolutionary Bot, Beijing 100093, Peoples R China 2.Univ Chinese Acad Sci, Beijing 100049, Peoples R China
推荐引用方式 GB/T 7714	Wang, Juanjuan,Wang, Yang,Lu, Tianfeng,et al. An Efficient and Universal Protoplast Isolation Protocol Suitable for Transient Gene Expression Analysis and Single-Cell RNA Sequencing[J]. INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES,2022,23(7).
APA	Wang, Juanjuan.,Wang, Yang.,Lu, Tianfeng.,Yang, Xia.,Liu, Jing.,...&Wang, Yinzheng.(2022).An Efficient and Universal Protoplast Isolation Protocol Suitable for Transient Gene Expression Analysis and Single-Cell RNA Sequencing.INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES,23(7).
MLA	Wang, Juanjuan,et al."An Efficient and Universal Protoplast Isolation Protocol Suitable for Transient Gene Expression Analysis and Single-Cell RNA Sequencing".INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES 23.7(2022).