TY - JOUR
T1 - Quantitative, super-resolution localization of small RNAs with sRNA-PAINT
AU - Huang, Kun
AU - Demirci, Feray
AU - Batish, Mona
AU - Treible, Wayne
AU - Meyers, Blake C.
AU - Caplan, Jeffrey L.
N1 - Funding Information:
US NSF Plant Genome Research Program [1649424, 1611853, 1754097]; Meyers and Caplan labs for their support; Joanna Friesner for editorial assistance. Funding for open access charge: NSF EAGER [1822293].
Publisher Copyright:
© 2020 The Author(s) 2020. Published by Oxford University Press on behalf of Nucleic Acids Research.
PY - 2020/9/18
Y1 - 2020/9/18
N2 - Small RNAs are non-coding RNAs that play important roles in the lives of both animals and plants. They are 21- to 24-nt in length and ∼10 nm in size. Their small size and high diversity have made it challenging to develop detection methods that have sufficient resolution and specificity to multiplex and quantify. We created a method, sRNA-PAINT, for the detection of small RNAs with 20 nm resolution by combining the super-resolution method, DNA-based points accumulation in nanoscale topography (DNA-PAINT), and the specificity of locked nucleic acid (LNA) probes for the in situ detection of multiple small RNAs. The method relies on designing probes to target small RNAs that combine DNA oligonucleotides (oligos) for PAINT with LNA-containing oligos for hybridization; therefore, we developed an online tool called 'Vetting & Analysis of RNA for in situ Hybridization probes' (VARNISH) for probe design. Our method utilizes advances in DNA-PAINT methodologies, including qPAINT for quantification, and Exchange-PAINT for multiplexing. We demonstrated these capabilities of sRNA-PAINT by detecting and quantifying small RNAs in different cell layers of early developmental stage maize anthers that are important for male sexual reproduction.
AB - Small RNAs are non-coding RNAs that play important roles in the lives of both animals and plants. They are 21- to 24-nt in length and ∼10 nm in size. Their small size and high diversity have made it challenging to develop detection methods that have sufficient resolution and specificity to multiplex and quantify. We created a method, sRNA-PAINT, for the detection of small RNAs with 20 nm resolution by combining the super-resolution method, DNA-based points accumulation in nanoscale topography (DNA-PAINT), and the specificity of locked nucleic acid (LNA) probes for the in situ detection of multiple small RNAs. The method relies on designing probes to target small RNAs that combine DNA oligonucleotides (oligos) for PAINT with LNA-containing oligos for hybridization; therefore, we developed an online tool called 'Vetting & Analysis of RNA for in situ Hybridization probes' (VARNISH) for probe design. Our method utilizes advances in DNA-PAINT methodologies, including qPAINT for quantification, and Exchange-PAINT for multiplexing. We demonstrated these capabilities of sRNA-PAINT by detecting and quantifying small RNAs in different cell layers of early developmental stage maize anthers that are important for male sexual reproduction.
UR - http://www.scopus.com/inward/record.url?scp=85091263901&partnerID=8YFLogxK
U2 - 10.1093/nar/gkaa623
DO - 10.1093/nar/gkaa623
M3 - Article
C2 - 32716042
AN - SCOPUS:85091263901
SN - 0305-1048
VL - 48
SP - E96
JO - Nucleic Acids Research
JF - Nucleic Acids Research
IS - 16
ER -