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| Name | Authors | DOI | Group |
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Vitamin Solution for ESAW Media for Marine Phytoplankton Resource Report Resource Website |
Ashley Humphrey | DOI:10.17504/protocols.io.gdcbs2w | Protist Research to Optimize Tools in Genetics (PROT-G), The Aquatic Microbial Ecology Research Group - AMERG (The Buchan, Zinser and Wilhelm labs) | Stock vitamin solution for ESAW Media for Marine Phytoplankton | University of Tennessee, Knoxville | 1 | 2016 | Ashley Humphrey 2016. Vitamin Solution for ESAW Media for Marine Phytoplankton. protocols.io https://dx.doi.org/10.17504/protocols.io.gdcbs2w | 2021-04-15 09:16:01 | |||
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CRISPR-Directed Mitotic Recombination Enables Genetic Mapping Without Crosses Resource Report Resource Website |
Meru J. Sadhu, Joshua S. Bloom, Laura Day, Leonid Kruglyak | DOI:10.17504/protocols.io.hu7b6zn | Protist Research to Optimize Tools in Genetics (PROT-G) | Please contact Dr. Steven Wilhelm ([email protected]) for additional information regarding this protocol. | , , , | http://science.sciencemag.org/content/352/6289/1113.full | 1 | 2017 | Meru J. Sadhu, Joshua S. Bloom, Laura Day, Leonid Kruglyak 2017. CRISPR-Directed Mitotic Recombination Enables Genetic Mapping Without Crosses. protocols.io https://dx.doi.org/10.17504/protocols.io.hu7b6zn | 2021-04-15 09:16:01 | ||
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Low Gel Temperature Agarose (LGTA) Media Resource Report Resource Website |
G Jason Smith, April Woods | DOI:10.17504/protocols.io.gmnbu5e | Protist Research to Optimize Tools in Genetics (PROT-G), Environmental Biotechnology Lab | Use for solid phase culture and chemical selection of Pseudo-nitzschia spp and other recalcitrant diatoms | EBL - Moss Landing Marine Laboratories, EBL - Moss Landing Marine Laboratories | 2 | 2016 | G Jason Smith, April Woods 2016. Low Gel Temperature Agarose (LGTA) Media. protocols.io https://dx.doi.org/10.17504/protocols.io.gmnbu5e | 2021-04-15 09:16:02 | |||
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Stable tranfection of unicellular relative of animals, Corallochytrium limacisporum, using Lonza Nucleofector Resource Report Resource Website |
Aleksandra Kozyczkowska | DOI:10.17504/protocols.io.r5ud86w | Protist Research to Optimize Tools in Genetics (PROT-G), Multicellgenomelab, Emerging Organisms for Biology | Institut de Biologia Evolutiva (UPF-CSIC) Barcelona | 1 | 2018 | Aleksandra Kozyczkowska 2018. Stable tranfection of unicellular relative of animals, Corallochytrium limacisporum, using Lonza Nucleofector. protocols.io https://dx.doi.org/10.17504/protocols.io.r5ud86w | 2021-04-15 09:16:05 | ||||
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Modified Qiagen PCR purification (no gel extraction) with MinElute spin column. Resource Report Resource Website |
Daniel Richter | DOI:10.17504/protocols.io.ivece3e | Ecology of Marine Plankton (ECOMAP) team - Roscoff, Protist Research to Optimize Tools in Genetics (PROT-G) | Richter, Daniel J and Fozouni, Parinaz and Eisen, Michael and King, Nicole. Gene family innovation, conservation and loss on the animal stem lineage. 2018;7:e34226 https://doi.org/10.7554/eLife.34226 | https://elifesciences.org/articles/34226 | 1 | 2017 | Daniel Richter 2017. Modified Qiagen PCR purification (no gel extraction) with MinElute spin column.. protocols.io https://dx.doi.org/10.17504/protocols.io.ivece3e | 2021-04-15 09:16:05 | |||
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Cell deposition and drug sensitivity assay Resource Report Resource Website |
Lev Tsypin, Aaron Turkewitz | DOI:10.17504/protocols.io.336gqre | Protist Research to Optimize Tools in Genetics (PROT-G) | This protocol describes how to deposit several cells of S. robusta in droplets on a petri dish, allowing the experimenter to track the growth of approximately single cell isolates with high throughput. This was used to determine drug sensitivity.We found that in, liquid culture, S. robusta is sensitive to 10 ug/mL puromycin and 250 ug/mL glyphosate. One caveat to keep in mind is that S. robusta appears more tolerant to antibiotics when grown on agar plates, being insensitive to up to 20 ug/mL of puromycin, for example. We recommend selection in liquid culture. | California Institute of Technology, University of Chicago | 1 | 2019 | Lev Tsypin, Aaron Turkewitz 2019. Cell deposition and drug sensitivity assay. protocols.io https://dx.doi.org/10.17504/protocols.io.336gqre | 2021-04-15 09:16:08 | |||
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HA tag enables highly efficient detection of heterologous proteins in Phaeodactylum tricornutum (Pt) exconjugants Resource Report Resource Website |
Jernej Turnsek | DOI:10.17504/protocols.io.j7ncrme | Protist Research to Optimize Tools in Genetics (PROT-G) | Invitrogen's HA tag monoclonal antibody in combination with Western Breeze Anti-Mouse Chemiluminescent Western Blot Immunodetection Kit leads to no background Western blots with Pt lysates. HA tag thus represents a tag of choice for (1) efficient and unambioguous protein expression analysis, (2) immunofluorescence experiments and (3) various IP and pull-down experiments in Pt and possibly other diatoms as well as other marine microeukaryotes. | University of California, Berkeley | 2 | 2017 | Jernej Turnsek 2017. HA tag enables highly efficient detection of heterologous proteins in Phaeodactylum tricornutum (Pt) exconjugants. protocols.io https://dx.doi.org/10.17504/protocols.io.j7ncrme | 2021-04-15 09:16:08 | |||
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Quantitative (q)PCR and Differential Expression Analysis Resource Report Resource Website |
Jeffrey Lewis, Amanda Scholes | DOI:10.17504/protocols.io.bbgpijvn | Protist Research to Optimize Tools in Genetics (PROT-G), Lewis Lab | A generalizable protocol for measuring relative changes in gene expression via qPCR (cDNA synthesis, qPCR primer optimization, and qPCR analysis), with specific optimization for the budding yeast Saccharomyces cerevisiae. | Scholes AN, Lewis JA, Comparison of RNA isolation methods on RNA-Seq: implications for differential expression and meta-analyses. BMC Genomics doi: 10.1186/s12864-020-6673-2 | University of Arkansas - Fayetteville, University of Arkansas - Fayetteville | https://doi.org/10.1186/s12864-020-6673-2 | 1 | 2020 | Jeffrey Lewis, Amanda Scholes 2020. Quantitative (q)PCR and Differential Expression Analysis. protocols.io https://dx.doi.org/10.17504/protocols.io.bbgpijvn | 2021-04-15 09:16:10 | |
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FlowCam Standard Operating Procedure Resource Report Resource Website |
Jacob Harris | DOI:10.17504/protocols.io.f2tbqen | Protist Research to Optimize Tools in Genetics (PROT-G), Alverson Lab | The FlowCam can be used to take quick cell density measurements of liquid samples. | 2 | 2016 | Jacob Harris 2016. FlowCam Standard Operating Procedure. protocols.io https://dx.doi.org/10.17504/protocols.io.f2tbqen | 2021-04-15 09:16:11 | ||||
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Growth of Dunaliella salina on artificial seawater. Resource Report Resource Website |
Larry Klobutcher | DOI:10.17504/protocols.io.gwpbxdn | Protist Research to Optimize Tools in Genetics (PROT-G) | University of Connecticut, School of medicine | 1 | 2017 | Larry Klobutcher 2017. Growth of Dunaliella salina on artificial seawater.. protocols.io https://dx.doi.org/10.17504/protocols.io.gwpbxdn | 2021-04-15 09:16:11 | ||||
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Golden Gate plasmids used for transfection of Perkinsus marinus Resource Report Resource Website |
Elin Einarsson | DOI:10.17504/protocols.io.37egrje | Protist Research to Optimize Tools in Genetics (PROT-G) | We have created plasmids for transfection of the parasite Perkinsus marinus using Golden Gate cloning. We have explored the possibility to either FACS sort (GFP/mCherry fused to MOE membrane protein) or using drug selection (bleomycin, blasticidin S or puromycin) to enrich transfected cells. | Waller Lab, Department of Biochemistry, University of Cambridge | 1 | 2019 | Elin Einarsson 2019. Golden Gate plasmids used for transfection of Perkinsus marinus. protocols.io https://dx.doi.org/10.17504/protocols.io.37egrje | 2021-04-15 09:16:12 | |||
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Bodo saltans SSU-GFP-Neo cassette Resource Report Resource Website |
Fatma Gomaa, Virginia Edgcomb, Peter Girguis, Zhuhong li, Roberto Docampo | DOI:10.17504/protocols.io.s5meg46 | Protist Research to Optimize Tools in Genetics (PROT-G) | This plasmid is designed to integrate within the small subunit ribosomal RNA gene region of B. saltans. It carries 1 kb of the B. saltans small subunit ribosomal RNA gene. To construct this cassette, we first sequenced the complete ribosomal operon of B. saltans, which has a total length of ̴9 kb. The structure of the ribosomal operon of B. saltans is similar in the closely related kinetoplastida Leishmania, where the ribosomal RNA genes are arranged in head- to-tail repeats (Figure 4b and Yan et al., Molecular and biochemical parasitology 103, no. 2 (1999): 197-210). For the cassette construction, we used only a 1 kb region of the 18S ribosomal RNA gene, which is represented in the schematic as Homologous Recombination Region 1 (HR1) and Homologous Recombination Region 2 (HR2) each, of which is 500 bp . The GFP and Neo genes are located in the middle of the construct and their expression is controlled by the presence of the B. saltans tubulin intergenic region (Bs TUB IGR) at the 3’ and 5’ ends of each gene. As the transcription in B. saltans is polycistronic, the presence of the tubulin intergenic region is necessary for trans-splicing and polyadenylation. | Harvard University, Woods Hole Oceanographic Institution, Harvard University, University of Georgia, University of Georgia | 1 | 2018 | Fatma Gomaa, Virginia Edgcomb, Peter Girguis, Zhuhong li, Roberto Docampo 2018. Bodo saltans SSU-GFP-Neo cassette. protocols.io https://dx.doi.org/10.17504/protocols.io.s5meg46 | 2021-04-15 09:16:17 | |||
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Submixture for Step1 of Labyrinthula growth medium Resource Report Resource Website |
Janina Brakel | DOI:10.17504/protocols.io.ehmbb46 | Protist Research to Optimize Tools in Genetics (PROT-G), Collier Lab | 1 | 2016 | Janina Brakel 2016. Submixture for Step1 of Labyrinthula growth medium. protocols.io https://dx.doi.org/10.17504/protocols.io.ehmbb46 | 2021-04-15 09:16:17 | |||||
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Small-scale silencing experiment in vegetative Euplotes crassus (provisional) Resource Report Resource Website |
Rachele Cesaroni | DOI:10.17504/protocols.io.2asgaee | Protist Research to Optimize Tools in Genetics (PROT-G) | Universität Bern | 1 | 2019 | Rachele Cesaroni 2019. Small-scale silencing experiment in vegetative Euplotes crassus (provisional). protocols.io https://dx.doi.org/10.17504/protocols.io.2asgaee | 2021-04-15 09:16:17 | ||||
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Euplotes crassus transformation using FuGene HD Transfection Reagent as vehicle Resource Report Resource Website |
RACHELE CESARONI | DOI:10.17504/protocols.io.g43byyn | Protist Research to Optimize Tools in Genetics (PROT-G) | University of Bern, Institute of Biology | 5 | 2017 | RACHELE CESARONI 2017. Euplotes crassus transformation using FuGene HD Transfection Reagent as vehicle. protocols.io https://dx.doi.org/10.17504/protocols.io.g43byyn | 2021-04-15 09:16:20 | ||||
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How to culture corallochytrean Corallochytrium limacisporum, a unicellular relative to animals Resource Report Resource Website |
Maria Rubio-Brotons | DOI:10.17504/protocols.io.gqmbvu6 | Protist Research to Optimize Tools in Genetics (PROT-G), Multicellgenomelab | IBE CSIC-UPF | 1 | 2016 | Maria Rubio-Brotons 2016. How to culture corallochytrean Corallochytrium limacisporum, a unicellular relative to animals. protocols.io https://dx.doi.org/10.17504/protocols.io.gqmbvu6 | 2021-04-15 09:16:22 | ||||
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Electroporation of natural communities in sea water Resource Report Resource Website |
Andrés Ramírez, Albane Ruaud, Fernan Federici, Peter von Dassow | DOI:10.17504/protocols.io.hpnb5me | Protist Research to Optimize Tools in Genetics (PROT-G) | This protocol has been optimized for electroporation of natural communities in coastal surface waters. As natural communities may vary in different places we recomend to use this protocol as a starting point and re-optimize according to the results. Guidelines are provided. | Instituto Milenio de Oceanografía, Instituto Milenio de Oceanografía, Instituto Milenio de Oceanografía, Instituto Milenio de Oceanografía | 2 | 2017 | Andrés Ramírez, Albane Ruaud, Fernan Federici, Peter von Dassow 2017. Electroporation of natural communities in sea water. protocols.io https://dx.doi.org/10.17504/protocols.io.hpnb5me | 2021-04-15 09:16:15 | |||
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Oyster parasite Perkinsus marinus transformation using Amaxa electroporator and non-proprietary electroporation buffer Resource Report Resource Website |
Imen Lassadi | DOI:10.17504/protocols.io.gv9bw96 | Protist Research to Optimize Tools in Genetics (PROT-G) | Faktorová D, Nisbet RER, Robledo JAF, Casacuberta E, Sudek L, Allen AE, Ares M, Aresté C, Balestreri C, Barbrook AC, Beardslee P, Bender S, Booth DS, Bouget F, Bowler C, Breglia SA, Brownlee C, Burger G, Cerutti H, Cesaroni R, Chiurillo MA, Clemente T, Coles DB, Collier JL, Cooney EC, Coyne K, Docampo R, Dupont CL, Edgcomb V, Einarsson E, Elustondo PA, Federici F, Freire-Beneitez V, Freyria NJ, Fukuda K, García PA, Girguis PR, Gomaa F, Gornik SG, Guo J, Hampl V, Hanawa Y, Haro-Contreras ER, Hehenberger E, Highfield A, Hirakawa Y, Hopes A, Howe CJ, Hu I, Ibañez J, Irwin NAT, Ishii Y, Janowicz NE, Jones AC, Kachale A, Fujimura-Kamada K, Kaur B, Kaye JZ, Kazana E, Keeling PJ, King N, Klobutcher LA, Lander N, Lassadi I, Li Z, Lin S, Lozano J, Luan F, Maruyama S, Matute T, Miceli C, Minagawa J, Moosburner M, Najle SR, Nanjappa D, Nimmo IC, Noble L, Vanclová AMGN, Nowacki M, Nuñez I, Pain A, Piersanti A, Pucciarelli S, Pyrih J, Rest JS, Rius M, Robertson D, Ruaud A, Ruiz-Trillo I, Sigg MA, Silver PA, Slamovits CH, Smith GJ, Sprecher BN, Stern R, Swart EC, Tsaousis AD, Tsypin L, Turkewitz A, Turnšek J, Valach M, Vergé V, Dassow Pv, Haar Tvd, Waller RF, Wang L, Wen X, Wheeler G, Woods A, Zhang H, Mock T, Worden AZ, Lukeš J, Genetic tool development in marine protists: emerging model organisms for experimental cell biology. Nature Methods 17(5). doi: 10.1038/s41592-020-0796-x | Department of Biochemistry, Cambridge university, UK. | https://doi.org/10.1038/s41592-020-0796-x | 1 | 2017 | Imen Lassadi 2017. Oyster parasite Perkinsus marinus transformation using Amaxa electroporator and non-proprietary electroporation buffer. protocols.io https://dx.doi.org/10.17504/protocols.io.gv9bw96 | 2021-04-15 09:16:21 | ||
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Electroporation of Heterosigma akashiwo Resource Report Resource Website |
Deepak Nanjappa and Kathryn Coyne | DOI:10.17504/protocols.io.hjmb4k6 | Protist Research to Optimize Tools in Genetics (PROT-G) | This protocol was developed for transformation of Heterosigma akashiwo by electroporation. This species does not grow on solid medium, so transformants are grown "in bulk" and should not be considered clonal. We have had limited success with this protocol, which may need to be further optimized. | University of Delaware | 2 | 2017 | Deepak Nanjappa and Kathryn Coyne 2017. Electroporation of Heterosigma akashiwo. protocols.io https://dx.doi.org/10.17504/protocols.io.hjmb4k6 | 2021-04-15 09:16:14 | |||
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Modified Bacterial Conjugation Protocol For Pseudo-nitzschia multiseries Resource Report Resource Website |
April Woods, G Jason Smith | DOI:10.17504/protocols.io.4pzgvp6 | Protist Research to Optimize Tools in Genetics (PROT-G), Environmental Biotechnology Lab | This is a modified version of 'Conjugation of Thalassiosira pseudonana', publishedby J. Turnsek dx.doi.org/10.17504/protocols.io.f55bq86 .The protocol was modified to enhance viability in the diatom Pseudo-nitzschia multiseries. This version was used to transform Pseudo-nitzschia multiseries isolate 15091C3 with an episomal plasmid derived from pPtPUC3 to express a egfp gene under control of the P. multiseries actin promoter and termination domains. PLASMID INFOpPmAGFPC10 EPI : A P. multiseries derived expression cassette under control of P. multiseries ca 1000 bp of actin promoter and termination domains. Pm actPs::eGFP::actTs | Faktorová D, Nisbet RER, Robledo JAF, Casacuberta E, Sudek L, Allen AE, Ares M, Aresté C, Balestreri C, Barbrook AC, Beardslee P, Bender S, Booth DS, Bouget F, Bowler C, Breglia SA, Brownlee C, Burger G, Cerutti H, Cesaroni R, Chiurillo MA, Clemente T, Coles DB, Collier JL, Cooney EC, Coyne K, Docampo R, Dupont CL, Edgcomb V, Einarsson E, Elustondo PA, Federici F, Freire-Beneitez V, Freyria NJ, Fukuda K, García PA, Girguis PR, Gomaa F, Gornik SG, Guo J, Hampl V, Hanawa Y, Haro-Contreras ER, Hehenberger E, Highfield A, Hirakawa Y, Hopes A, Howe CJ, Hu I, Ibañez J, Irwin NAT, Ishii Y, Janowicz NE, Jones AC, Kachale A, Fujimura-Kamada K, Kaur B, Kaye JZ, Kazana E, Keeling PJ, King N, Klobutcher LA, Lander N, Lassadi I, Li Z, Lin S, Lozano J, Luan F, Maruyama S, Matute T, Miceli C, Minagawa J, Moosburner M, Najle SR, Nanjappa D, Nimmo IC, Noble L, Vanclová AMGN, Nowacki M, Nuñez I, Pain A, Piersanti A, Pucciarelli S, Pyrih J, Rest JS, Rius M, Robertson D, Ruaud A, Ruiz-Trillo I, Sigg MA, Silver PA, Slamovits CH, Smith GJ, Sprecher BN, Stern R, Swart EC, Tsaousis AD, Tsypin L, Turkewitz A, Turnšek J, Valach M, Vergé V, Dassow Pv, Haar Tvd, Waller RF, Wang L, Wen X, Wheeler G, Woods A, Zhang H, Mock T, Worden AZ, Lukeš J, Genetic tool development in marine protists: emerging model organisms for experimental cell biology. Nature Methods 17(5). doi: 10.1038/s41592-020-0796-x | Moss Landing Marine Labs, Moss Landing Marine Labs | https://doi.org/10.1038/s41592-020-0796-x | 1 | 2019 | April Woods, G Jason Smith 2019. Modified Bacterial Conjugation Protocol For Pseudo-nitzschia multiseries. protocols.io https://dx.doi.org/10.17504/protocols.io.4pzgvp6 | 2021-04-15 09:16:28 |
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