Space and time-resolved gene expression experiments on cultured mammalian cells by a single-cell electroporation microarray.
Single-cell experiments represent the next frontier for biochemical and gene expression research. Although bulk-scale methods averaging populations of cells have been traditionally used to investigate cellular behavior, they mask individual cell features and can lead to misleading or insufficient biological results. We report on a single-cell electroporation microarray enabling the transfection of pre-selected individual cells at different sites within the same culture (space-resolved), at arbitrarily chosen time points and even sequentially to the same cells (time-resolved). Delivery of impermeant molecules by single-cell electroporation was first proven to be finely tunable by acting on the electroporation protocol and then optimized for transfection of nucleic acids into Chinese Hamster Ovary (CHO-K1) cells. We focused on DNA oligonucleotides (ODNs), short interfering RNAs (siRNAs), and DNA plasmid vectors, thus providing a versatile and easy-to-use platform for time-resolved gene expression experiments in single mammalian cells.
Vassanelli
S
S
University of Padova, Department of Human Anatomy and Physiology, Section of Physiology, via Marzolo 3 - 35131, Padova, Italy. stefano.vassanelli@unipd.it
Bandiera
L
L
Borgo
M
M
Cellere
G
G
Santoni
L
L
Bersani
C
C
Salamon
M
M
Zaccolo
M
M
Lorenzelli
L
L
Girardi
S
S
Maschietto
M
M
Dal Maschio
M
M
Paccagnella
A
A
D000818:Animals; D016466:CHO Cells; D002478:Cells, Cultured; D006224:Cricetinae; D003412:Cricetulus; D018274:Electroporation / Q000379:methods*; D005786:Gene Expression Regulation; D020868:Gene Silencing; D049452:Green Fluorescent Proteins / Q000378:metabolism; D042541:Intracellular Space / Q000378:metabolism; D008839:Microelectrodes; D009696:Nucleic Acids / Q000378:metabolism; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods*; D009841:Oligonucleotides / Q000378:metabolism; D034741:RNA, Small Interfering / Q000378:metabolism; D013997:Time Factors; D014162:Transfection
D009696:Nucleic Acids; D009841:Oligonucleotides; D034741:RNA, Small Interfering; C527701:enhanced cyan fluorescent protein; D049452:Green Fluorescent Proteins