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Authors: Janina Brakel
Group: Protist Research to Optimize Tools in Genetics (PROT-G), Collier Lab
Proper citation: Janina Brakel 2016. Labyrinthula growth medium. protocols.io dx.doi.org/10.17504/protocols.io.ehkbb4w Copy
Authors: Elena Casacuberta, Cristina Aresté, Omaya Dudin , Elena Casacuberta
Group: Protist Research to Optimize Tools in Genetics (PROT-G), Multicellgenomelab, Emerging Organisms for Biology
Summary: Testing electroporation on Spaheroforma artica with two different electroporatorsNeon (invitrogen)Lonza (nucleofector)
Proper citation: Elena Casacuberta, Cristina Aresté, Omaya Dudin , Elena Casacuberta 2019. Unsucessful atemps on transfromation of the Icthyosporean Sphaeroforma artica. protocols.io dx.doi.org/10.17504/protocols.io.z6ef9be Copy
Authors: Oskar Johansson, Adrian Clarke
Group: Protist Research to Optimize Tools in Genetics (PROT-G)
Summary: Mapping of transformed DNA within mutated diatoms. A stepwise specific PCR reaction that allow for identification of genome location of transformed fragments such as antibiotic resistance genes or GFP constructs. Degenerate primer 1: 5' - GCCGGACTTAGCGAGCA TGTGTAGTACCTTGCCG ATCCTNASATANSATANT TC - 3'Degenerate primer 2: 5' - GCCGGACTTAGCGAGCA TGTGTAGTACCTTGCCG ATCCCNTSABGNACYTN CTG - 3'Degenerate primer 3: 5' - GCCGGACTTAGCGAGCA TGTGTAGTACCTTGCCG ATCCNGACGARWGANA WGAC - 3'Degenerate primer 4: 5' - GCCGGACTTAGCGAGCA TGTGTAGTACCTTGCCG ATCCTAHATGDAGKACN TAC - 3'Specific primer 1: 5' - GCCGGACTTAGCGAGCA TGTGTAG - 3'Specific primer 2: 5' - CATGTGTAGTACCTTGC CGATCC - 3'Internal primers must be designed with regard to each construct that are to be mapped. They should not overlap but instead cover consecutive stretches of known DNA sequence of the transformed construct.
Proper citation: Oskar Johansson, Adrian Clarke 2019. Genomic mapping of transformed DNA fragments. protocols.io dx.doi.org/10.17504/protocols.io.yq2fvye Copy
Authors: Lawrence A. Klobutcher
Group: Protist Research to Optimize Tools in Genetics (PROT-G)
Summary: The electroporation procedure and conditions described allow for the entry of DAPI into the cell, resulting in nuclear staining. We have not yet been successful in using these conditions for transformation of Euplotes crassus, but only a couple of constructs have been tried to date.
Proper citation: Lawrence A. Klobutcher 2017. Electroporation of Euplotes crassus: conditions that allow for the uptake of DAPI.. protocols.io dx.doi.org/10.17504/protocols.io.gwmbxc6 Copy
Authors: G Jason Smith
Group: Protist Research to Optimize Tools in Genetics (PROT-G), Environmental Biotechnology Lab
Summary: This is a modification of the origianl Chelex 100 extraction described by Walsh, Metzger and Higuchi (1991 BioTechniques 10(4):506). Adding PVPP facilitates working with environmental samples as well as pure cultures hi in phenolics or other contaminants.
Proper citation: G Jason Smith 2017. Simple DNA Extraction for Phytoplankton Using Chelex 100. protocols.io dx.doi.org/10.17504/protocols.io.h58b89w Copy
Authors: David Booth
Group: King Lab, Protist Research to Optimize Tools in Genetics (PROT-G)
Proper citation: David Booth 2018. Protocol G: Preparation of reporter DNA. protocols.io dx.doi.org/10.17504/protocols.io.h8yb9xw Copy
Authors: Angela Piersanti
Group: Protist Research to Optimize Tools in Genetics (PROT-G)
Proper citation: Angela Piersanti 2019. Transfection by Electroporation in Euplotes crassus. protocols.io dx.doi.org/10.17504/protocols.io.2a9gah6 Copy
Authors: Fatma Gomma, Paulo A. Garcia, Jennifer Delaney, Peter R. Girguis, Cullen R. Buie, Virginia P. Edgcomb
Group: Protist Research to Optimize Tools in Genetics (PROT-G)
Summary: The process of developing transient transfection protocols in P. caudatus involved initial testing and determination of proper electroporation buffers and parameters (voltage strength, pulse duration and number) to maximize cell viability. Our results suggest that the type of electroporation buffer is critical for maintaining cell viability throughout the experiment and is essential for determining the optimum electric field range.
Proper citation: Fatma Gomma, Paulo A. Garcia, Jennifer Delaney, Peter R. Girguis, Cullen R. Buie, Virginia P. Edgcomb 2017. Parabodo caudatus post-electroporation cell viability quantification using exponential decay electroporation. protocols.io dx.doi.org/10.17504/protocols.io.hmhb436 Copy
Authors: Angela Piersanti
Group: Protist Research to Optimize Tools in Genetics (PROT-G)
Proper citation: Angela Piersanti 2017. Euplotes transfection using Lipofectamine 3000 (provisional). protocols.io dx.doi.org/10.17504/protocols.io.hhub36w Copy
Authors: Jernej Turnsek, Chris Dupont
Group: Protist Research to Optimize Tools in Genetics (PROT-G)
Summary: Here we present an efficient way to grow Tp colonies inside a 0.25% superclean agar matrix. This protocol could possibly be applicable to other marine microeukaryotes that are problematic to grow on fully solid support.
Proper citation: Jernej Turnsek, Chris Dupont 2017. Pour plating of Thalassiosira pseudonana (Tp). protocols.io dx.doi.org/10.17504/protocols.io.i6qchdw Copy
Authors: Mariana Rius, Jackie Collier
Group: Protist Research to Optimize Tools in Genetics (PROT-G), Collier Lab
Summary: Electroporation strategy used in the transformation of Aurantiochytrium limacinum (ATCC MYA-1381; Stramenopile/ Heterokont, Thraustochytrid). This protocol is following the guidance of Ono et al. 2011. US Patent # 7,888,123 B2.
Proper citation: Mariana Rius, Jackie Collier 2018. Electroporation of Aurantiochytrium limacinum (ATCC MYA-1381). protocols.io dx.doi.org/10.17504/protocols.io.qjcduiw Copy
Authors: Alle Lie
Group: Protist Research to Optimize Tools in Genetics (PROT-G), Caron Lab - Protistan Ecology
Proper citation: Alle Lie 2016. Making heat-killed bacteria for feeding axenic phagotrophic protists. protocols.io dx.doi.org/10.17504/protocols.io.ewkbfcw Copy
Authors: Anastasios Tsaousis
Group: Protist Research to Optimize Tools in Genetics (PROT-G)
Proper citation: Anastasios Tsaousis 2017. Transfected Naegleria Fluorescence Microscopy. protocols.io dx.doi.org/10.17504/protocols.io.hpvb5n6 Copy
Authors: Meru J. Sadhu, Joshua S. Bloom, Laura Day, Leonid Kruglyak
Group: Protist Research to Optimize Tools in Genetics (PROT-G)
Summary: Please contact Dr. Steven Wilhelm ([email protected]) for additional information regarding this protocol.
Proper citation: Meru J. Sadhu, Joshua S. Bloom, Laura Day, Leonid Kruglyak 2017. CRISPR-Directed Mitotic Recombination Enables Genetic Mapping Without Crosses. protocols.io dx.doi.org/10.17504/protocols.io.hu7b6zn Copy
Authors: Aleksandra Kozyczkowska
Group: Protist Research to Optimize Tools in Genetics (PROT-G), Multicellgenomelab, Emerging Organisms for Biology
Proper citation: Aleksandra Kozyczkowska 2018. Stable tranfection of unicellular relative of animals, Corallochytrium limacisporum, using Lonza Nucleofector. protocols.io dx.doi.org/10.17504/protocols.io.r5ud86w Copy
Authors: Ashley Humphrey
Group: Protist Research to Optimize Tools in Genetics (PROT-G), The Aquatic Microbial Ecology Research Group - AMERG (The Buchan, Zinser and Wilhelm labs)
Summary: Stock vitamin solution for ESAW Media for Marine Phytoplankton
Proper citation: Ashley Humphrey 2016. Vitamin Solution for ESAW Media for Marine Phytoplankton. protocols.io dx.doi.org/10.17504/protocols.io.gdcbs2w Copy
Authors: G Jason Smith, April Woods
Group: Protist Research to Optimize Tools in Genetics (PROT-G), Environmental Biotechnology Lab
Summary: Use for solid phase culture and chemical selection of Pseudo-nitzschia spp and other recalcitrant diatoms
Proper citation: G Jason Smith, April Woods 2016. Low Gel Temperature Agarose (LGTA) Media. protocols.io dx.doi.org/10.17504/protocols.io.gmnbu5e Copy
Authors: Ashley Humphrey
Group: Protist Research to Optimize Tools in Genetics (PROT-G), The Aquatic Microbial Ecology Research Group - AMERG (The Buchan, Zinser and Wilhelm labs)
Summary: Stock vitamin solution for ESAW Media for Marine Phytoplankton
Proper citation: Ashley Humphrey 2016. Vitamin Solution for ESAW Media for Marine Phytoplankton. protocols.io dx.doi.org/10.17504/protocols.io.ge9bth6 Copy
Authors: Gina Filloramo
Group: Protist Research to Optimize Tools in Genetics (PROT-G)
Summary: Successful transformation of Phaeodactylum tricornutum (CMP632) by electroporation was achieved based on a modified version of Zhang & Hu (2014).
Proper citation: Gina Filloramo 2018. Transformation of Phaeodactylum tricornutum by electroporation. protocols.io dx.doi.org/10.17504/protocols.io.s4cegsw Copy
Authors: Daniel Richter
Group: Ecology of Marine Plankton (ECOMAP) team - Roscoff, Protist Research to Optimize Tools in Genetics (PROT-G)
Proper citation: Daniel Richter 2017. Modified Qiagen PCR purification (no gel extraction) with MinElute spin column.. protocols.io dx.doi.org/10.17504/protocols.io.ivece3e Copy
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