11. Embryos transferred to 19
C before 10% epiboly will not
survive. Embryos transferred to temperatures below 19
C at
any point will die.
12. In our hands, maximal intensity under 561 nm laser is attained
about 1 h after photoconversion. For this reason, embryos are
illuminated with 405 nm laser and then incubated for 1 h
before further processing either to dissociation or imaging.
Also, photoconverted Dendra2 is stable for about 36 h after
photoconversion.
Acknowledgments
We are in debt to Nicolas Daudet for his help and support. The
Linker lab has been supported by grants from the Wellcome Trust,
Royal Society, and the Medical Research council. SH and LB are
funded by ICM P09-015-F, DAAD 57220037 & 57168868,
CORFO 16CTTS-66390, Fondecyt 1181823, FONDEF
19I10334.
References
1. Stramer BM, Dunn GA (2015) Cells on film—
the past and future of cinemicroscopy. J Cell Sci
128:9–13.
https://doi.org/10.1242/jcs.
165019
2. BFI. Cheese Mites (1903) | BFI National
Archive. n.d.
3. Meyers JR (2018) Zebrafish: development of a
vertebrate model organism. Curr Protoc
Essent Lab Technol 16:e19. https://doi.org/
10.1002/cpet.19
4. Brabletz T, Kalluri R, Nieto MA, Weinberg RA
(2018) EMT in cancer. Nat Rev Cancer
5. Delloye-Bourgeois C, Castellani V (2019)
Hijacking of embryonic programs by neural
crest-derived neuroblastoma: from physiological migration to metastatic dissemination.
Front Mol Neurosci 12. https://doi.org/10.
3389/fnmol.2019.00052
6. Le Douarin NM, Kalcheim C (1999) The neural crest, 2nd edn. Cambridge University Press,
Cambridge
7. Raible DW, Wood A, Hodsdon W, Henion PD,
Weston JA, Eisen JS (1992) Segregation and
early dispersal of neural crest cells in the embryonic zebrafish. Dev Dyn 195:29–42. https://
doi.org/10.1002/aja.1001950104
8. Richardson
J,
Gauert
A,
Briones
Montecinos L, Fanlo L, Alhashem ZM, Assar
R et al (2016) Leader cells define directionality
of trunk, but not cranial, neural crest cell
migration. Cell Rep 15:2076–2088. https://
doi.org/10.1016/j.celrep.2016.04.067
9. Duffy JB (2002) GAL4 system indrosophila: a
fly geneticist’s swiss army knife. Genesis
34:1–15.
https://doi.org/10.1002/gene.
10150
10. Distel M, Wullimann MF, Ko ¨ster RW (2009)
Optimized Gal4 genetics for permanent gene
expression mapping in zebrafish. Proc Natl
Acad Sci U S A 106:13365–13370. https://
doi.org/10.1073/pnas.0903060106
11. Collins RT, Linker C, Lewis J (2010) MAZe: a
tool for mosaic analysis of gene function in
zebrafish. Nat Methods 7:219–223. https://
doi.org/10.1038/nmeth.1423
12. Gorelik R, Gautreau A (2014) Quantitative
and unbiased analysis of directional persistence
in cell migration. Nat Protoc 9:1931–1943.
https://doi.org/10.1038/nprot.2014.131
13. Kwan KM, Fujimoto E, Grabher C, Mangum
BD, Hardy ME, Campbell DS et al (2007) The
Tol2kit: a multisite gateway-based construction kit forTol2 transposon transgenesis constructs. Dev Dyn 236:3088–3099. https://
doi.org/10.1002/dvdy.21343
14. Chong-Morrison V, Simoes FC, Senanayake U,
Carroll DS, Riley PR, Sauka-Spengler T (2018)
Re-purposing Ac/Ds transgenic system for
CRISPR/dCas9 modulation of enhancers and
non-coding RNAs in zebrafish. BioRxiv
Tools to Study Neural Crest Migration in Zebrafish
105
C before 10% epiboly will not
survive. Embryos transferred to temperatures below 19
C at
any point will die.
12. In our hands, maximal intensity under 561 nm laser is attained
about 1 h after photoconversion. For this reason, embryos are
illuminated with 405 nm laser and then incubated for 1 h
before further processing either to dissociation or imaging.
Also, photoconverted Dendra2 is stable for about 36 h after
photoconversion.
Acknowledgments
We are in debt to Nicolas Daudet for his help and support. The
Linker lab has been supported by grants from the Wellcome Trust,
Royal Society, and the Medical Research council. SH and LB are
funded by ICM P09-015-F, DAAD 57220037 & 57168868,
CORFO 16CTTS-66390, Fondecyt 1181823, FONDEF
19I10334.
References
1. Stramer BM, Dunn GA (2015) Cells on film—
the past and future of cinemicroscopy. J Cell Sci
128:9–13.
https://doi.org/10.1242/jcs.
165019
2. BFI. Cheese Mites (1903) | BFI National
Archive. n.d.
3. Meyers JR (2018) Zebrafish: development of a
vertebrate model organism. Curr Protoc
Essent Lab Technol 16:e19. https://doi.org/
10.1002/cpet.19
4. Brabletz T, Kalluri R, Nieto MA, Weinberg RA
(2018) EMT in cancer. Nat Rev Cancer
5. Delloye-Bourgeois C, Castellani V (2019)
Hijacking of embryonic programs by neural
crest-derived neuroblastoma: from physiological migration to metastatic dissemination.
Front Mol Neurosci 12. https://doi.org/10.
3389/fnmol.2019.00052
6. Le Douarin NM, Kalcheim C (1999) The neural crest, 2nd edn. Cambridge University Press,
Cambridge
7. Raible DW, Wood A, Hodsdon W, Henion PD,
Weston JA, Eisen JS (1992) Segregation and
early dispersal of neural crest cells in the embryonic zebrafish. Dev Dyn 195:29–42. https://
doi.org/10.1002/aja.1001950104
8. Richardson
J,
Gauert
A,
Briones
Montecinos L, Fanlo L, Alhashem ZM, Assar
R et al (2016) Leader cells define directionality
of trunk, but not cranial, neural crest cell
migration. Cell Rep 15:2076–2088. https://
doi.org/10.1016/j.celrep.2016.04.067
9. Duffy JB (2002) GAL4 system indrosophila: a
fly geneticist’s swiss army knife. Genesis
34:1–15.
https://doi.org/10.1002/gene.
10150
10. Distel M, Wullimann MF, Ko ¨ster RW (2009)
Optimized Gal4 genetics for permanent gene
expression mapping in zebrafish. Proc Natl
Acad Sci U S A 106:13365–13370. https://
doi.org/10.1073/pnas.0903060106
11. Collins RT, Linker C, Lewis J (2010) MAZe: a
tool for mosaic analysis of gene function in
zebrafish. Nat Methods 7:219–223. https://
doi.org/10.1038/nmeth.1423
12. Gorelik R, Gautreau A (2014) Quantitative
and unbiased analysis of directional persistence
in cell migration. Nat Protoc 9:1931–1943.
https://doi.org/10.1038/nprot.2014.131
13. Kwan KM, Fujimoto E, Grabher C, Mangum
BD, Hardy ME, Campbell DS et al (2007) The
Tol2kit: a multisite gateway-based construction kit forTol2 transposon transgenesis constructs. Dev Dyn 236:3088–3099. https://
doi.org/10.1002/dvdy.21343
14. Chong-Morrison V, Simoes FC, Senanayake U,
Carroll DS, Riley PR, Sauka-Spengler T (2018)
Re-purposing Ac/Ds transgenic system for
CRISPR/dCas9 modulation of enhancers and
non-coding RNAs in zebrafish. BioRxiv
Tools to Study Neural Crest Migration in Zebrafish
105