TY - JOUR
T1 - An optimized dissociation protocol for FACS-based isolation of rare cell types from Caenorhabditis elegans L1 larvae
AU - Fernandes Póvoa, Euclides E
AU - Ebbing, Annabel L P
AU - Betist, Marco C
AU - van der Veen, Christa
AU - Korswagen, Hendrik C
N1 - Funding Information:
We would like to thank Stefan van der Elst from the Flow Cytometry Core facility (Hubrecht Institute) for technical support. We are also grateful to David M. Miller, III and Rebecca McWhirter for helpful advices during the optimization of this protocol. This work is part of the research program (14NOISE01) of the Foundation for Fundamental Research on Matter (FOM), which is financially supported by the Netherlands Organization for Scientific Research (NWO). Some strains were provided by the CGC, which is funded by NIH Office of Research Infrastructure Programs (P40 OD010440).
Funding Information:
We would like to thank Stefan van der Elst from the Flow Cytometry Core facility (Hubrecht Institute) for technical support. We are also grateful to David M. Miller, III and Rebecca McWhirter for helpful advices during the optimization of this protocol. This work is part of the research program ( 14NOISE01 ) of the Foundation for Fundamental Research on Matter (FOM) , which is financially supported by the Netherlands Organization for Scientific Research (NWO) . Some strains were provided by the CGC, which is funded by NIH Office of Research Infrastructure Programs ( P40 OD010440 ).
Publisher Copyright:
© 2020 The Author(s)
PY - 2020
Y1 - 2020
N2 - Single-cell isolation and transcriptomic analysis of a specific cell type or tissue offers the possibility of studying cell function and heterogeneity in time-dependent processes with remarkable resolution. The reduced tissue complexity and highly stereotyped development of Caenorhabditis elegans, combined with an extensive genetic toolbox and the ease of growing large tightly synchronized populations makes it an exceptional model organism for the application of such approaches. However, the difficulty to dissociate and isolate single cells from larval stages has been a major constraint to this kind of studies. Here, we describe an improved protocol for dissociation and preparation of single cell suspensions from developmentally synchronized populations of C. elegans L1 larvae. Our protocol has been empirically optimized to allow efficient FACS-based purification of large number of single cells from rare cell types, for subsequent extraction and sequencing of their mRNA.
AB - Single-cell isolation and transcriptomic analysis of a specific cell type or tissue offers the possibility of studying cell function and heterogeneity in time-dependent processes with remarkable resolution. The reduced tissue complexity and highly stereotyped development of Caenorhabditis elegans, combined with an extensive genetic toolbox and the ease of growing large tightly synchronized populations makes it an exceptional model organism for the application of such approaches. However, the difficulty to dissociate and isolate single cells from larval stages has been a major constraint to this kind of studies. Here, we describe an improved protocol for dissociation and preparation of single cell suspensions from developmentally synchronized populations of C. elegans L1 larvae. Our protocol has been empirically optimized to allow efficient FACS-based purification of large number of single cells from rare cell types, for subsequent extraction and sequencing of their mRNA.
KW - C. elegans
KW - L1 larvae dissociation
KW - Single cell isolation
UR - http://www.scopus.com/inward/record.url?scp=85085663339&partnerID=8YFLogxK
U2 - 10.1016/j.mex.2020.100922
DO - 10.1016/j.mex.2020.100922
M3 - Article
C2 - 32509539
SN - 2215-0161
VL - 7
SP - 1
EP - 6
JO - MethodsX
JF - MethodsX
M1 - 100922
ER -