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Wheat Root-dip Inoculation with Fusarium graminearum and Assessment of Root Rot Disease Severity

Featured protocol,  Authors: Qing Wang
Qing Wang Affiliation: Department of Plant Breeding, IFZ Research Centre for Biosystems, Land Use and Nutrition, Justus Liebig University, Giessen, Germany
Bio-protocol author page: a4241
 and Sven Gottwald
Sven GottwaldAffiliation: Department of Plant Breeding, IFZ Research Centre for Biosystems, Land Use and Nutrition, Justus Liebig University, Giessen, Germany
For correspondence: sv.gottwald@t-online.de
Bio-protocol author page: a4240
date: 3/20/2017, 127 views, 0 Q&A
DOI: https://doi.org/10.21769/BioProtoc.2189.

Brief version appeared in Mol Plant Microbe Interact, Dec 2015
Fusarium graminearum is one of the most common and potent fungal pathogens of wheat (Triticum aestivum) and other cereals, known for causing devastating yield losses and mycotoxin contaminations of food and feed. The pathogen is mainly considered as a paradigm for the floral disease Fusarium head blight, while its ability to colonize wheat plants via root infection has been examined recently. F. graminearum has a unique infection strategy which comprises complex, specialized structures and processes. Root colonisation negatively affects plant development and leads to systemic plant invasion by tissue-adapted fungal strategies. The pathosystem wheat root - F. graminearum makes available an array of research areas, such as (i) the relatively unknown root interactions with a necrotrophic pathogen; (ii) genes and pathways contributing to (overall) Fusarium resistance; (iii) induced systemic (whole-plant) resistance; (iv) pathogenic strategies in a variety of host tissues; and (v) age-related changes in the single-genotype responses to seedling and adult plant (root/spike) infection. The presented Fusarium root rot bioassay allows for efficient infection of wheat roots, evaluation of disease severity and progress as well as statistical analysis of disease dynamics.

Xylem Sap Extraction Method from Hop Plants

Featured protocol,  Authors: Marko Flajšman*
Marko FlajšmanAffiliation: Department of Agronomy, Biotechnical Faculty, University of Ljubljana, Ljubljana, Slovenia
Bio-protocol author page: a4167
Stanislav Mandelc*
Stanislav MandelcAffiliation: Department of Agronomy, Biotechnical Faculty, University of Ljubljana, Ljubljana, Slovenia
Bio-protocol author page: a4168
Sebastjan Radišek*
Sebastjan RadišekAffiliation: Slovenian Institute of Hop Research and Brewing, Zalec, Slovenia
Bio-protocol author page: a4169
 and Branka Javornik
Branka JavornikAffiliation: Department of Agronomy, Biotechnical Faculty, University of Ljubljana, Ljubljana, Slovenia
For correspondence: branka.javornik@bf.uni-lj.si
Bio-protocol author page: a4170
 (*contributed equally to this work) date: 3/20/2017, 121 views, 0 Q&A
DOI: https://doi.org/10.21769/BioProtoc.2172.

Brief version appeared in Mol Plant Microbe Interact, May 2016
Verticillium wilt is one of the most important diseases on hop that significantly influence continuation of production on affected areas. It is caused by the soil borne vascular pathogen Verticillium nonalfalfae, which infects plants through the roots and then advances through the vascular (xylem) system. During infection, V. nonalfalfae secretes many different virulence factors. Xylem sap of infected plants is therefore a rich source for investigating the molecules that are involved in molecular interactions of Verticillium – hop plants. This protocol provides instructions on how to infect hop plants with V. nonalfalfae artificially and how to obtain xylem sap from hop plants.

Pathogenicity Assay of Verticillium nonalfalfae on Hop Plants

Featured protocol,  Authors: Marko Flajšman*
Marko FlajšmanAffiliation: Department of Agronomy, Biotechnical Faculty, University of Ljubljana, Ljubljana, Slovenia
Bio-protocol author page: a4167
Sebastjan Radišek*
Sebastjan RadišekAffiliation: Slovenian Institute of Hop Research and Brewing, Zalec, Slovenia
Bio-protocol author page: a4169
 and Branka Javornik
Branka JavornikAffiliation: Department of Agronomy, Biotechnical Faculty, University of Ljubljana, Ljubljana, Slovenia
For correspondence: branka.javornik@bf.uni-lj.si
Bio-protocol author page: a4170
 (*contributed equally to this work) date: 3/20/2017, 106 views, 0 Q&A
DOI: https://doi.org/10.21769/BioProtoc.2171.

Brief version appeared in Mol Plant Microbe Interact, May 2016
Verticillium nonalfalfae is a soil-borne plant pathogen that infects its hosts through roots. It spreads in the plant’s xylem and causes wilt disease symptoms by secreting different virulence factors. Hop (Humulus lupulus) is a primary host of V. nonalfalfae, so it is used as a model plant for studying this phytopathogenic fungus. Artificial infections of hop plants and disease scoring are prerequisites for studying the pathogen’s virulence/pathogenicity and its interaction with hop plants. In this protocol we describe the root dipping inoculation method for conducting pathogenicity assay of V. nonalfalfae on hop plants.

Polyethylene Glycol-mediated Transformation of Drechmeria coniospora

Featured protocol,  Authors: Le D. He
Le D. HeAffiliation: Aix Marseille Univ, CNRS, INSERM, CIML, Centre d’Immunologie de Marseille-Luminy, Marseille, France
Bio-protocol author page: a4195
 and Jonathan J. Ewbank
Jonathan J. EwbankAffiliation: Aix Marseille Univ, CNRS, INSERM, CIML, Centre d’Immunologie de Marseille-Luminy, Marseille, France
For correspondence: ewbank@ciml.univ-mrs.fr
Bio-protocol author page: a4196
date: 3/5/2017, 193 views, 0 Q&A
DOI: https://doi.org/10.21769/BioProtoc.2157.

Brief version appeared in PLoS Genet, May 2016
Drechmeria coniospora is a nematophagous fungus and potential biocontrol agent. It belongs to the Ascomycota. It is related to Hirsutella minnesotensis, another nematophagous fungus but, phylogenetically, it is currently closest to the truffle parasite Tolypocladium ophioglossoides. Together with its natural host, Caenorhabditis elegans, it is used to study host-pathogen interactions. Here, we report a polyethylene glycol-mediated transformation method (Turgeon et al., 2010; Ochman et al., 1988) for this fungus. The protocol can be used to generate both knock-in or knock-out strains (Lebrigand et al., 2016).

Determination of Elemental Concentrations in Lichens Using ICP-AES/MS

Featured protocol,  Authors: Liang-Cheng Zhao
Liang-Cheng ZhaoAffiliation: Hebei Geological Laboratory, Baoding, China
Bio-protocol author page: a4076
Li Wang
Li WangAffiliation: Hebei Geological Laboratory, Baoding, China
Bio-protocol author page: a4077
Yun-Jun Jiang
Yun-Jun JiangAffiliation: Hebei Geological Laboratory, Baoding, China
Bio-protocol author page: a4078
Yan-Qiao Hu
Yan-Qiao HuAffiliation: Hebei Geological Laboratory, Baoding, China
Bio-protocol author page: a4122
Chong-Ying Xu
Chong-Ying XuAffiliation: Hebei Geological Laboratory, Baoding, China
Bio-protocol author page: a4080
Lei Wang
Lei WangAffiliation: Hebei Geological Laboratory, Baoding, China
Bio-protocol author page: a4081
Xing Li
Xing Li Affiliation: Hebei Geological Laboratory, Baoding, China
Bio-protocol author page: a4082
Li Wei
Li WeiAffiliation: Hebei Geological Laboratory, Baoding, China
Bio-protocol author page: a4083
Xiu-Ping Guo
Xiu-Ping GuoAffiliation: Hebei Geological Laboratory, Baoding, China
Bio-protocol author page: a4084
Ai-Qin Liu
Ai-Qin LiuAffiliation: Hebei Geological Laboratory, Baoding, China
For correspondence: laq217510@sina.com
Bio-protocol author page: a4086
 and Hua-Jie Liu
Hua-Jie LiuAffiliation: College of Life Sciences, Hebei University, Baoding, China
For correspondence: liuhuajie@foxmail.com
Bio-protocol author page: a4087
date: 3/5/2017, 161 views, 0 Q&A
DOI: https://doi.org/10.21769/BioProtoc.2165.

Brief version appeared in Sci Rep, Apr 2016
Lichens are good biomonitors for air pollution because of their high enrichment capability of atmospheric chemical elements. To monitor atmospheric element deposition using lichens, it is important to obtain information on the multi-element concentrations in lichen thalli. Because of serious air pollution, elemental concentrations in thalli of lichens from North China (especially Inner Mongolia, Hebei, Shanxi and Henan province) are often higher than those from other regions, therefore highlighting the necessity to optimize ICP-AES/MS (Inductively coupled plasma-atomic emission spectroscopy/mass spectrometry) for analyzing lichen element content. Based on the high elemental concentrations in the lichen samples, and the differences in the sensitivity and detection limits between ICP-MS and ICP-AES, we propose a protocol for analyzing 31 elements in lichens using ICP-AES/MS. Twenty-two elements (Cd, Ce, Co, Cr, Cs, Cu, K, La, Mo, Na, Ni, Pb, Rb, Sb, Sc, Sm, Sr, Tb, Th, Tl, V and Zn) can be identified by using microwave digestion- ICP-MS, and 9 elements (Al, Ba, Ca, Fe, Mg, Mn, P, S and Ti) by using ashing-alkali fusion digestion- ICP-AES.

Chitin Extraction and Content Measurement in Magnaporthe oryzae

Featured protocol,  Authors: Xinyu Liu
Xinyu LiuAffiliation: Department of Plant Pathology, College of Plant Protection, Nanjing Agricultural University, and Key Laboratory of Integrated Management of Crop Diseases and Pests, Ministry of Education, Nanjing, China
Bio-protocol author page: a4123
 and Zhengguang Zhang
Zhengguang ZhangAffiliation: Department of Plant Pathology, College of Plant Protection, Nanjing Agricultural University, and Key Laboratory of Integrated Management of Crop Diseases and Pests, Ministry of Education, Nanjing, China
For correspondence: zhgzhang@njau.edu.cn
Bio-protocol author page: a4208
date: 3/5/2017, 202 views, 0 Q&A
DOI: https://doi.org/10.21769/BioProtoc.2164.

Brief version appeared in Mol Plant Microbe Interact, Jun 2016
Chitin is a linear polysaccharide composed of β (1→4)-linked N-acetylglucosamine (GlcNAc) residues. In fungi, chitin is an important component of the cell wall. Here, we provide a protocol to measure the chitin content of fungal cells using Magnaporthe oryzae as an example.

Wheat Root-dip Inoculation with Fusarium graminearum and Assessment of Root Rot Disease Severity

Authors: Qing Wang
Qing Wang Affiliation: Department of Plant Breeding, IFZ Research Centre for Biosystems, Land Use and Nutrition, Justus Liebig University, Giessen, Germany
Bio-protocol author page: a4241
 and Sven Gottwald
Sven GottwaldAffiliation: Department of Plant Breeding, IFZ Research Centre for Biosystems, Land Use and Nutrition, Justus Liebig University, Giessen, Germany
For correspondence: sv.gottwald@t-online.de
Bio-protocol author page: a4240
date: 3/20/2017, 127 views, 0 Q&A
DOI: https://doi.org/10.21769/BioProtoc.2189.

[Abstract] Fusarium graminearum is one of the most common and potent fungal pathogens of wheat (Triticum aestivum) and other cereals, known for causing devastating yield losses and mycotoxin contaminations of food and feed. The pathogen is mainly considered as a paradigm for the floral disease Fusarium head blight, ...

Xylem Sap Extraction Method from Hop Plants

Authors: Marko Flajšman*
Marko FlajšmanAffiliation: Department of Agronomy, Biotechnical Faculty, University of Ljubljana, Ljubljana, Slovenia
Bio-protocol author page: a4167
Stanislav Mandelc*
Stanislav MandelcAffiliation: Department of Agronomy, Biotechnical Faculty, University of Ljubljana, Ljubljana, Slovenia
Bio-protocol author page: a4168
Sebastjan Radišek*
Sebastjan RadišekAffiliation: Slovenian Institute of Hop Research and Brewing, Zalec, Slovenia
Bio-protocol author page: a4169
 and Branka Javornik
Branka JavornikAffiliation: Department of Agronomy, Biotechnical Faculty, University of Ljubljana, Ljubljana, Slovenia
For correspondence: branka.javornik@bf.uni-lj.si
Bio-protocol author page: a4170
 (*contributed equally to this work) date: 3/20/2017, 121 views, 0 Q&A
DOI: https://doi.org/10.21769/BioProtoc.2172.

[Abstract] Verticillium wilt is one of the most important diseases on hop that significantly influence continuation of production on affected areas. It is caused by the soil borne vascular pathogen Verticillium nonalfalfae, which infects plants through the roots and then advances through the vascular (xylem) system. ...

Pathogenicity Assay of Verticillium nonalfalfae on Hop Plants

Authors: Marko Flajšman*
Marko FlajšmanAffiliation: Department of Agronomy, Biotechnical Faculty, University of Ljubljana, Ljubljana, Slovenia
Bio-protocol author page: a4167
Sebastjan Radišek*
Sebastjan RadišekAffiliation: Slovenian Institute of Hop Research and Brewing, Zalec, Slovenia
Bio-protocol author page: a4169
 and Branka Javornik
Branka JavornikAffiliation: Department of Agronomy, Biotechnical Faculty, University of Ljubljana, Ljubljana, Slovenia
For correspondence: branka.javornik@bf.uni-lj.si
Bio-protocol author page: a4170
 (*contributed equally to this work) date: 3/20/2017, 106 views, 0 Q&A
DOI: https://doi.org/10.21769/BioProtoc.2171.

[Abstract] Verticillium nonalfalfae is a soil-borne plant pathogen that infects its hosts through roots. It spreads in the plant’s xylem and causes wilt disease symptoms by secreting different virulence factors. Hop (Humulus lupulus) is a primary host of V. nonalfalfae, so it is used as a model plant for studying ...

Polyethylene Glycol-mediated Transformation of Drechmeria coniospora

Authors: Le D. He
Le D. HeAffiliation: Aix Marseille Univ, CNRS, INSERM, CIML, Centre d’Immunologie de Marseille-Luminy, Marseille, France
Bio-protocol author page: a4195
 and Jonathan J. Ewbank
Jonathan J. EwbankAffiliation: Aix Marseille Univ, CNRS, INSERM, CIML, Centre d’Immunologie de Marseille-Luminy, Marseille, France
For correspondence: ewbank@ciml.univ-mrs.fr
Bio-protocol author page: a4196
date: 3/5/2017, 193 views, 0 Q&A
DOI: https://doi.org/10.21769/BioProtoc.2157.

[Abstract] Drechmeria coniospora is a nematophagous fungus and potential biocontrol agent. It belongs to the Ascomycota. It is related to Hirsutella minnesotensis, another nematophagous fungus but, phylogenetically, it is currently closest to the truffle parasite Tolypocladium ophioglossoides. Together with its ...

Determination of Elemental Concentrations in Lichens Using ICP-AES/MS

Authors: Liang-Cheng Zhao
Liang-Cheng ZhaoAffiliation: Hebei Geological Laboratory, Baoding, China
Bio-protocol author page: a4076
Li Wang
Li WangAffiliation: Hebei Geological Laboratory, Baoding, China
Bio-protocol author page: a4077
Yun-Jun Jiang
Yun-Jun JiangAffiliation: Hebei Geological Laboratory, Baoding, China
Bio-protocol author page: a4078
Yan-Qiao Hu
Yan-Qiao HuAffiliation: Hebei Geological Laboratory, Baoding, China
Bio-protocol author page: a4122
Chong-Ying Xu
Chong-Ying XuAffiliation: Hebei Geological Laboratory, Baoding, China
Bio-protocol author page: a4080
Lei Wang
Lei WangAffiliation: Hebei Geological Laboratory, Baoding, China
Bio-protocol author page: a4081
Xing Li
Xing Li Affiliation: Hebei Geological Laboratory, Baoding, China
Bio-protocol author page: a4082
Li Wei
Li WeiAffiliation: Hebei Geological Laboratory, Baoding, China
Bio-protocol author page: a4083
Xiu-Ping Guo
Xiu-Ping GuoAffiliation: Hebei Geological Laboratory, Baoding, China
Bio-protocol author page: a4084
Ai-Qin Liu
Ai-Qin LiuAffiliation: Hebei Geological Laboratory, Baoding, China
For correspondence: laq217510@sina.com
Bio-protocol author page: a4086
 and Hua-Jie Liu
Hua-Jie LiuAffiliation: College of Life Sciences, Hebei University, Baoding, China
For correspondence: liuhuajie@foxmail.com
Bio-protocol author page: a4087
date: 3/5/2017, 161 views, 0 Q&A
DOI: https://doi.org/10.21769/BioProtoc.2165.

[Abstract] Lichens are good biomonitors for air pollution because of their high enrichment capability of atmospheric chemical elements. To monitor atmospheric element deposition using lichens, it is important to obtain information on the multi-element concentrations in lichen thalli. Because of serious air pollution, ...

Chitin Extraction and Content Measurement in Magnaporthe oryzae

Authors: Xinyu Liu
Xinyu LiuAffiliation: Department of Plant Pathology, College of Plant Protection, Nanjing Agricultural University, and Key Laboratory of Integrated Management of Crop Diseases and Pests, Ministry of Education, Nanjing, China
Bio-protocol author page: a4123
 and Zhengguang Zhang
Zhengguang ZhangAffiliation: Department of Plant Pathology, College of Plant Protection, Nanjing Agricultural University, and Key Laboratory of Integrated Management of Crop Diseases and Pests, Ministry of Education, Nanjing, China
For correspondence: zhgzhang@njau.edu.cn
Bio-protocol author page: a4208
date: 3/5/2017, 202 views, 0 Q&A
DOI: https://doi.org/10.21769/BioProtoc.2164.

[Abstract] Chitin is a linear polysaccharide composed of β (1→4)-linked N-acetylglucosamine (GlcNAc) residues. In fungi, chitin is an important component of the cell wall. Here, we provide a protocol to measure the chitin content of fungal cells using Magnaporthe oryzae as an example. ...

Penetration Assays, Fungal Recovery and Pathogenicity Assays for Verticillium dahliae

Authors: Yun-Long Zhao
Yun-Long ZhaoAffiliation: State Key Laboratory of Plant Genomics, Institute of Microbiology, Chinese Academy of Sciences, Beijing, China
For correspondence: zyl714@126.com
Bio-protocol author page: a4092
Tao Zhang
Tao Zhang Affiliation: State Key Laboratory of Plant Genomics, Institute of Microbiology, Chinese Academy of Sciences, Beijing, China
Bio-protocol author page: a4093
 and Hui-Shan Guo
Hui-Shan GuoAffiliation: State Key Laboratory of Plant Genomics, Institute of Microbiology, Chinese Academy of Sciences, Beijing, China
Bio-protocol author page: a2721
date: 2/20/2017, 300 views, 0 Q&A
DOI: https://doi.org/10.21769/BioProtoc.2133.

[Abstract] Verticillium dahliae is a soil-borne phytopathogenic fungus that infects host roots and proliferates in vascular tissues. The great loss of economically important crop caused by V. dahliae has raised worldwide concern, however, little is known about the mechanism of its pathogenicity (Klosterman et ...

In vitro Histone H3 Cleavage Assay for Yeast and Chicken Liver H3 Protease

Authors: Sakshi Chauhan
Sakshi ChauhanAffiliation: Laboratory of Chromatin Biology, Department of Biological Sciences, Indian Institute of Science Education and Research, Bhopal, India
Bio-protocol author page: a3946
Gajendra Kumar Azad
Gajendra Kumar AzadAffiliation 1: Laboratory of Chromatin Biology, Department of Biological Sciences, Indian Institute of Science Education and Research, Bhopal, India
Affiliation 2: Department of Genetics, Institute of Life Sciences, Hebrew University of Jerusalem, Jerusalem, Israel
Bio-protocol author page: a3947
 and Raghuvir Singh Tomar
Raghuvir Singh TomarAffiliation: Laboratory of Chromatin Biology, Department of Biological Sciences, Indian Institute of Science Education and Research, Bhopal, India
For correspondence: rst@iiserb.ac.in
Bio-protocol author page: a3948
date: 1/5/2017, 594 views, 0 Q&A
DOI: https://doi.org/10.21769/BioProtoc.2085.

[Abstract] Histone proteins are subjected to a wide array of reversible and irreversible post-translational modifications (PTMs) (Bannister and Kouzarides, 2011; Azad and Tomar, 2014). The PTMs on histones are known to regulate chromatin structure and function. Histones are irreversibly modified by proteolytic ...

Plant Tissue Trypan Blue Staining During Phytopathogen Infection

Authors: Nuria Fernández-Bautista
Nuria Fernández-BautistaAffiliation: Centro de Biotecnología y Genómica de Plantas, Universidad Politécnica de Madrid (UPM) – Instituto Nacional de Investigación y Tecnología Agraria y Alimentaria (INIA), Parque Científico y Tecnológico, UPM, Campus de Montegancedo, Ctra M-40, km 38, 28223, Pozuelo de Alarcón (Madrid), Spain
Bio-protocol author page: a3925
José Alfonso Domínguez-Núñez
José Alfonso Domínguez-NúñezAffiliation: Departamento de Sistemas y Recursos Naturales, Escuela Técnica Superior de Ingeniería de Montes, Forestal y del Medio Natural, Ciudad Universitaria s/n, 28040, Madrid, Spain
Bio-protocol author page: a3926
M. Mar Castellano Moreno
M. Mar Castellano MorenoAffiliation: Centro de Biotecnología y Genómica de Plantas, Universidad Politécnica de Madrid (UPM) – Instituto Nacional de Investigación y Tecnología Agraria y Alimentaria (INIA), Parque Científico y Tecnológico, UPM, Campus de Montegancedo, Ctra M-40, km 38, 28223, Pozuelo de Alarcón (Madrid), Spain
Bio-protocol author page: a3927
 and Marta Berrocal-Lobo
Marta Berrocal-LoboAffiliation 1: Centro de Biotecnología y Genómica de Plantas, Universidad Politécnica de Madrid (UPM) – Instituto Nacional de Investigación y Tecnología Agraria y Alimentaria (INIA), Parque Científico y Tecnológico, UPM, Campus de Montegancedo, Ctra M-40, km 38, 28223, Pozuelo de Alarcón (Madrid), Spain
Affiliation 2: Departamento de Sistemas y Recursos Naturales, Escuela Técnica Superior de Ingeniería de Montes, Forestal y del Medio Natural, Ciudad Universitaria s/n, 28040, Madrid, Spain
For correspondence: m.berrocal@upm.es
Bio-protocol author page: a3928
date: 12/20/2016, 975 views, 0 Q&A
DOI: https://doi.org/10.21769/BioProtoc.2078.

[Abstract] In this protocol plant tissue is stained with trypan blue dye allowing the researcher to visualize cell death. Specifically this method avoids the use of the carcinogen compound chloral hydrate, making this classical method of staining safer and faster than never. The protocol is applied specifically ...

Assessment of Wheat Resistance to Fusarium graminearum by Automated Image Analysis of Detached Leaves Assay

Authors: Alexandre Perochon
Alexandre PerochonAffiliation: University College Dublin Earth Institute and School of Biology and Environmental Science, College of Science, University College Dublin, Belfield, Dublin, Ireland
For correspondence: alexandre.perochon@ucd.ie
Bio-protocol author page: a3890
 and Fiona M. Doohan
Fiona M. DoohanAffiliation: University College Dublin Earth Institute and School of Biology and Environmental Science, College of Science, University College Dublin, Belfield, Dublin, Ireland
Bio-protocol author page: a2308
date: 12/20/2016, 786 views, 0 Q&A
DOI: https://doi.org/10.21769/BioProtoc.2065.

[Abstract] Fusarium head blight (FHB) caused by Fusarium pathogens is a globally important cereal disease. To study Fusarium pathogenicity and host disease resistance, robust methods for disease assessment and quantification are needed. Here we describe the procedure of a detached leaves assay emphasizing the ...
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In vitro Protein Kinase Assay

Author: Yuehua Wei
Yuehua WeiAffiliation: Department of Pharmacology, Cancer Institute of New Jersey, UMDNJ Robert Wood Johnson Medical School, Piscataway, USA
For correspondence: weiyh.sjtu.edu@gmail.com
Bio-protocol author page: a49
date: 6/5/2012, 21457 views, 0 Q&A
DOI: https://doi.org/10.21769/BioProtoc.212.

[Abstract] This protocol will describe experimental procedures for an in vitro kinase assay of the yeast protein kinase Sch9. This protocol can be tailored to detect kinase activity of other yeast protein kinase....

[Bio101] Making Yeast Competent Cells and Yeast Cell Transformation

Author: Yongxian Lu
Yongxian LuAffiliation: Carnegie Institution for Science, Stanford University, Stanford, USA
For correspondence: yxlu@stanford.edu
Bio-protocol author page: a28
date: 7/20/2011, 20797 views, 2 Q&A
DOI: https://doi.org/10.21769/BioProtoc.96.

[Abstract] This is a quite simple but reliable protocol to make very high transformation efficiency yeast competent cells. By express your gene of interest, protein function can be studied in yeast cells....

[Bio101] Protocol for Whole Cell Lysis of Yeast

Author: Zongtian Tong
Zongtian TongAffiliation: Department of Cell Biology, Center for Metabolism and Obesity Research, Johns Hopkins School of Medicine, Baltimore, USA
For correspondence: tongzong@gmail.com
Bio-protocol author page: a14
date: 1/5/2011, 18414 views, 4 Q&A
DOI: https://doi.org/10.21769/BioProtoc.14.

[Abstract] This protocol describes how to perform lysis on whole yeast cell samples using NaOH. The lysed cells can then be used for downstream applications such as the extraction of total proteins. ...

Spot Assay for Yeast

Author: Zongtian Tong
Zongtian TongAffiliation: Department of Cell Biology, Center for Metabolism and Obesity Research, Johns Hopkins School of Medicine, Baltimore, USA
For correspondence: tongzong@gmail.com
Bio-protocol author page: a14
date: 1/5/2012, 17991 views, 3 Q&A
DOI: https://doi.org/10.21769/BioProtoc.16.

[Abstract] This protocol can be used to compare the cell growth rate of yeast under different growth conditions. It involves the serial dilution and spotting of yeast colonies....

[Bio101] Yeast Vacuole Staining with FM4-64

Author: Zongtian Tong
Zongtian TongAffiliation: Department of Cell Biology, Center for Metabolism and Obesity Research, Johns Hopkins School of Medicine, Baltimore, USA
For correspondence: tongzong@gmail.com
Bio-protocol author page: a14
date: 1/5/2011, 14336 views, 1 Q&A
DOI: https://doi.org/10.21769/BioProtoc.18.

[Abstract] The lipophilic probe, FM 4-64 does not fluoresce much in water but fluoresces strongly after binding to the outer plasma membrane, providing clear and distinguishable plasma membrane staining. The binding is rapid and reversible. In this protocol vacuoles in yeast cells are stained with the FM4-64 dye, ...

[Bio101] How to Use an Avestin Emulsiflex C3 Homogenizer to Disrupt Cells

Author: Zongtian Tong
Zongtian TongAffiliation: Department of Cell Biology, Center for Metabolism and Obesity Research, Johns Hopkins School of Medicine, Baltimore, USA
For correspondence: tongzong@gmail.com
Bio-protocol author page: a14
date: 1/5/2011, 11376 views, 1 Q&A
DOI: https://doi.org/10.21769/BioProtoc.11.

[Abstract] The EmulsiFlex-C3 homogenizer is powered by an electric motor. The pump does not require a compressor for it to run. This equipment can be used to disrupt cells at a large scale. The EmulsiFlex-C3 has a fixed flow-through capacity of 3 L/h. It has the ability to process samples as small as 10 ml. The ...

[Bio101] Large Scale Native Affinity Purifications of Solubilized Membrane Proteins from Yeast

Author: Zongtian Tong
Zongtian TongAffiliation: Department of Cell Biology, Center for Metabolism and Obesity Research, Johns Hopkins School of Medicin, Baltimore , USA
For correspondence: tongzong@gmail.com
Bio-protocol author page: a14
date: 1/5/2011, 10939 views, 0 Q&A
DOI: https://doi.org/10.21769/BioProtoc.12.

[Abstract] This protocol can be used to purify membrane proteins from yeast samples under native conditions at a large scale. This protocol has been developed primarily for FLAG-tagged proteins. This protocol can however be slightly modified and applied to other tags, such as GST or HA....

[Bio101] Small Scale Native Affinity Purifications of Solubilized Membrane Proteins from Yeast

Author: Zongtian Tong
Zongtian TongAffiliation: Department of Cell Biology, Center for Metabolism and Obesity Research, Johns Hopkins School of Medicine, Baltimore, USA
For correspondence: tongzong@gmail.com
Bio-protocol author page: a14
date: 1/5/2011, 10399 views, 1 Q&A
DOI: https://doi.org/10.21769/BioProtoc.15.

[Abstract] In this protocol, we show how to purify membrane proteins from yeast using affinity purification under native conditions at a small scale. ...

Illumina Sequencing Library Construction from ChIP DNA

Author: Wei Zheng
Wei ZhengAffiliation: Keck Biotech Services, Yale University, New Haven, USA
For correspondence: wei.zheng.madison@gmail.com
Bio-protocol author page: a10
date: 2/20/2012, 9877 views, 0 Q&A
DOI: https://doi.org/10.21769/BioProtoc.91.

[Abstract] The Illumina sequencing platform is very popular among next-generation sequencing platforms. However, the DNA sequencing library construction kit provided by Illumina is considerably expensive. The protocol described here can be used to construct high-quality sequencing libraries from chromatin immunoprecipitated ...

Co-immunoprecipitation in Yeast

Author: Olesya O. Panasenko
Olesya O. PanasenkoAffiliation: Department of Microbiology and Molecular Medicine, University of Geneva, Faculty of Medicine, CMU, Geneva, Switzerland
For correspondence: olesya.panasenko@unige.ch
Bio-protocol author page: a88
date: 8/20/2012, 9296 views, 0 Q&A
DOI: https://doi.org/10.21769/BioProtoc.250.

[Abstract] This protocol describes investigation of protein-protein interactions in baker yeast by co-immunoprecipitation (CoIP). CoIP is a technique to identify physiologically relevant protein-protein interactions in the cell. The interesting protein can be isolated out of solution using antibody that specifically ...
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