Name: genome-wide analysis of histone modifications distribution using the chromatin immunoprecipitation sequencing method in magnaporthe oryzae
Uploaded: 2021-06-02T14:00:00
Description: Watch this Scientific Journal Video about Genome-wide Analysis of Histone Modifications Distribution using the Chromatin Immunoprecipitation Sequencing Method in Magnaporthe oryzae at JoVE.com

This work was supported by National Natural Science Foundation of China (Grant no. 31871638), the Special Scientific Research Project of Beijing Agriculture University (YQ201603), the Scientific Project of Beijing Educational Committee (KM201610020005), the High-level scientific research cultivation project of BUA (GJB2021005).

1. Preparation of protoplasts from M. oryzae
<ol>
	<li>Prepare the oatmeal-tomato agar (OTA).
	<ol>
		<li>Weigh 30-50 g of oatmeal and boil it in 800 mL of water (ddH2O) for 20 min. Filter through two layers of gauze and take the filtrate.</li>
		<li>Pick ripe tomatoes and peel them. Squeeze the juice, and filter through two layers of gauze to collect 150 mL of the filtered juice.</li>
		<li>Mix all the tomato juice and the prepared oat filtrate thoroughly and add ddH2O up to 1000 mL.</li...

<ol>
	<li>Kornberg, R. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Chromatin+structure:+are+repeating+unit+of+histones+and+DNA.">Chromatin structure: are repeating unit of histones and DNA.</a> Science. 184 (4139), 868-871 (1974).</li><li>Luger, K., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Crystal+structure+of+the+nucleosomecore+particle+at+2.8+a+resolution.">Crystal structure of the nucleosomecore particle at 2.8 a resolution.</a> Nature. 389 (6648), 251-260 (1997).</li><li>Strathl, B. D., Allis, C. 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R., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Covalent+modifications+of+histones+during+development+and+disease+pathogenesis.">Covalent modifications of histones during development and disease pathogenesis.</a> Nature Structural and Molecular Biology. 14 (11), 1008-1016 (2007).</li><li>Shilatifard, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Molecular+implementation+and+physiological+roles+for+histone+H3+lysine+4+(H3K4)+methylation.">Molecular implementation and physiological roles for histone H3 lysine 4 (H3K4) methylation.</a> Current Opinion in Cell Biology. 20 (3), 341-348 (2008).</li><li>Berger, S. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+complex+language+of+chromatin+regulation+during+transcription.">The complex language of chromatin regulation during transcription.</a> Nature. 447 (7143), 407-412 (2007).</li><li>Bernstein, B. E., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+mammalian+epigenome.">The mammalian epigenome.</a> Cell. 128 (4), 669-681 (2007).</li><li>Weake, V. M., Workman, J. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Histone+ubiquitination:+triggering+gene+activity.">Histone ubiquitination: triggering gene activity.</a> Molecular Cell. 29 (6), 653-663 (2008).</li><li>Workman, J. L., Kingston, R. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Alteration+of+nucleosome+structure+as+a+mechanism+of+transcriptional+regulation.">Alteration of nucleosome structure as a mechanism of transcriptional regulation.</a> Annual Review of Biochemistry. 67 (1), 545-579 (2003).</li><li>Kouzarides, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Chromatin+modifications+and+their+function.">Chromatin modifications and their function.</a> Cell. 128 (4), 693-705 (2007).</li><li>Akhtar, J., More, P., Albrecht, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=ChIP-Seq+from+limited+starting+material+of+K562+cells+and+Drosophila+neuroblasts+using+tagmentation+assisted+fragmentation+approach.">ChIP-Seq from limited starting material of K562 cells and Drosophila neuroblasts using tagmentation assisted fragmentation approach.</a> Bio-protocol. 10 (4), 3520 (2020).</li><li>Steinhauser, S., Kurzawa, N., Eils, R., Herrmann, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+comprehensive+comparison+of+tools+of+differential+ChIP-seq+analysis.">A comprehensive comparison of tools of differential ChIP-seq analysis.</a> Briefings in Bioinformatics. 17 (6), 953-966 (2016).</li><li>Kieu, T., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=MoSET1+(histone+H3K4+methyltransferase+in+Magnaporthe+oryzae)+regulates+global+gene+expression+during+infection-related+morphogenesis.">MoSET1 (histone H3K4 methyltransferase in Magnaporthe oryzae) regulates global gene expression during infection-related morphogenesis.</a> Plos Genetics. 11 (7), 11005385 (2015).</li><li>Vu, B. V., Pham, K. T., Nakayashiki, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Substrate-induced+transcriptional+activation+of+the+MoCel7C+cellulase+gene+is+associated+with+methylation+of+histone+H3+at+lysine+4+in+the+rice+blast+fungus+Magnaporthe+oryzae.">Substrate-induced transcriptional activation of the MoCel7C cellulase gene is associated with methylation of histone H3 at lysine 4 in the rice blast fungus Magnaporthe oryzae.</a> Applied and Environmental Microbiology. 79 (21), 6823-6832 (2013).</li><li>Kazan, K., Gardiner, D. M., Manners, J. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=On+the+trail+of+a+cereal+killer:+recent+advances+in+Fusarium+graminearum+pathogenomics+and+host+resistance.">On the trail of a cereal killer: recent advances in Fusarium graminearum pathogenomics and host resistance.</a> Molecular Plant Pathology. 13 (4), 399-413 (2012).</li><li>Wang, G. H., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+AMT1+arginine+methyltransferase+gene+is+important+for+plant+infection+and+normal+hyphal+growth+in+Fusarium+graminearum.">The AMT1 arginine methyltransferase gene is important for plant infection and normal hyphal growth in Fusarium graminearum.</a> PLoS One. 7 (5), 38324 (2012).</li><li>Liu, Y., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Histone+H3K4+methylation+regulates+hyphal+growth,+secondary+metabolism+and+multiple+stress+responses+in+Fusarium+graminearum.">Histone H3K4 methylation regulates hyphal growth, secondary metabolism and multiple stress responses in Fusarium graminearum.</a> Environmental Microbiology. 17 (11), 4615-4630 (2016).</li><li>Zhang, M. Y., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+plant+infection+test:+spray+and+wound-mediated+inoculation+with+the+plant+pathogen+Magnaporthe+grisea.">The plant infection test: spray and wound-mediated inoculation with the plant pathogen Magnaporthe grisea.</a> Journal of Visualized Experiments. (138), e57675 (2018).</li><li>Connolly, L. R., Smith, K. M., Freitag, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+Fusarium+graminearum+histone+H3K27+methyltransferase+KMT6+regulates+development+and+expression+of+secondary+metabolite+gene+clusters.">The Fusarium graminearum histone H3K27 methyltransferase KMT6 regulates development and expression of secondary metabolite gene clusters.</a> PloS Genetics. 9 (10), 1003916 (2013).</li><li>Palmer, J. M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Loss+of+CclA,+required+for+histone+3+lysine+4+methylation,+decreases+growth+but+increases+secondary+metabolite+production+in+Aspergillus+fumigatus.">Loss of CclA, required for histone 3 lysine 4 methylation, decreases growth but increases secondary metabolite production in Aspergillus fumigatus.</a> PeerJ. 1, 4 (2013).</li><li>Ding, S. L., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+Tig1+histone+deacetylase+complex+regulates+infectious+growth+in+the+rice+blast+fungus+Magnaporthe+oryzae.">The Tig1 histone deacetylase complex regulates infectious growth in the rice blast fungus Magnaporthe oryzae.</a> The Plant Cell. 22 (7), 2495-2508 (2010).</li><li>Dean, R. A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+genome+sequence+of+the+rice+blast+fungus+Magnaporthe+grisea.">The genome sequence of the rice blast fungus Magnaporthe grisea.</a> Nature. 434 (7036), 980-986 (2005).</li><li>Allis, C. D., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=New+nomenclature+for+chromatin-modifying+enzymes.">New nomenclature for chromatin-modifying enzymes.</a> Cell. 131 (4), 633-636 (2007).</li><li>Shilatifard, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+COMPASS+family+of+histone+H3K4+methylases:+mechanisms+of+regulation+in+development+and+disease+pathogenesis.">The COMPASS family of histone H3K4 methylases: mechanisms of regulation in development and disease pathogenesis.</a> Annual Review of Biochemistry. 81, 65-95 (2012).</li><li>Zhou, S. D., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+COMPASS-like+complex+modulates+fungal+development+and+pathogenesis+by+regulating+H3K4me3-mediated+targeted+gene+expression+in+Magnaporthe+oryzae.">The COMPASS-like complex modulates fungal development and pathogenesis by regulating H3K4me3-mediated targeted gene expression in Magnaporthe oryzae.</a> Molecular Plant Pathology. 22 (4), 422-439 (2021).</li></ol>

Recently, ChIP-seq has become a widely used genomic analysis method for determining the binding sites of TFs or enrichment sites modified by specific histones. Compared to previous ChIP-seq technology, new ChIP-seq technology is highly sensitive and flexible. Results are provided in high resolution without negative effects, such as the noise signal caused by the non-specific hybridization of nucleic acids. Although this is a common gene expression analysis, many computational methods have been validated, and the complexi...

Epigenetics is a branch of genetic research that refers to the heritable change of gene expression without changing the nucleotide sequence of genes. An increasing number of studies have shown that epigenetic regulation plays an important role in the growth and development of eukaryotic cells, including chromatin that regulates and affects gene expression through the dynamic process of folding and assembly into higher-order structures1,2. Chromatin remodeling and covalent histone modification affect and regulate the function and structure of chromatin through the variation of chromatin polymers, thereby achieving the function of regulating gene expression3,4,5,6. Posttranslational modifications of histone include acetylation, phosphorylation, methylation, monoubiquitination, sumoylation, and ADP ribosylation7,8,9. Histone H3K4 methylation, particularly trimethylation, has been mapped to transcription start sites where it is associated with transcription replication, recombination, repair, and RNA processing in eukaryotes10,11.
ChIP-seq technology was introduced in 2007 and has become the experimental standard for the genome-wide analysis of transcriptional regulation and epigenetic mechanisms12,13. This method is suitable at the genome-wide scale and for obtaining histone or transcription factor interaction information, including DNA segments of DNA binding proteins. Any DNA sequences crosslinked to proteins of interest will coprecipitate as a part of the chromatin complex. New-generation sequencing techniques are also used to sequence 36-100 bp of DNA, which are then matched to the corresponding target genome.
In phytopathogenic fungi, research has recently begun to study how histone methylation modifications regulate their target genes in the process of pathogenicity. Some previous studies proved that the regulation of histone methylase-related genes is mainly reflected in gene silencing and catalyzing the production of Secondary Metabolites (SM). MoSet1 is the H3K4 methylase in M. oryzae. Knockout of this gene results in the complete deletion of H3K4me3 modification14. Compared with the wild-type strain, the expression of the gene MoCEL7C in the mutant is inhibited in the CMC-induced state and in the non-induced state (glucose or cellobiose), the expression of MoCEL7C increased15. In Fusarium graminearum, KMT6 can catalyze the methylation modification of H3K27me3, regulate the normal development of fungi, and help regulate the &#34;cryptic genome&#34; containing the SM gene cluster16,17,18,19. In 2013, Connolly reported that H3K9 and H3K27 methylation regulates the pathogenic process of fungi through secondary metabolites and effector factors that regulate the inhibition of target genes20. In Aspergillus, the modification of histones H3K4me2 and H3K4me3 is related to gene activation and plays an important role in controlling the chromatin level regulation of SM gene clusters21. In M. oryzae, Tig1 (homologous to Tig1 in yeast and mammals) is an HADC (histone deacetylase)22. Knockout of the Tig1 gene leads to the complete loss of pathogenicity and spore production ability in the null mutant. It is more sensitive to a peroxygen environment, which cannot produce infective hyphae22.
The rice blast caused by M. oryzae. is one of the most serious rice diseases in most rice-growing areas in the world19. Due to its representative infection process, M. oryzae is similar to the infection process of many important pathogenic fungi. As it can easily carry out molecular genetic operations, the fungus has become a model organism for studying fungal-plant interactions23. Blocking every step of the infection process of M. oryzae may result in unsuccessful infection. The morphological changes during the infection process are strictly regulated by the entire genome function and gene transcription. Among them, epigenetic modifications such as histone methylation play an essential role in the transcriptional regulation of functional genes24,25. However, so far, few studies have been done on the molecular mechanism of epigenetic modifications such as histone methylation and histone acetylation in the transcription of pathogenesis genes in M. oryzae. Therefore, further developing the epigenetic regulation mechanism of the rice blast fungus while researchinh the upstream and downstream regulatory network of these pathogenic related genes will help develop rice blast prevention and control strategies.
With the development of functional genomics such as ChIP-seq, especially in epigenetics, this high-throughput data acquisition method has accelerated research on chromosomes. Using the ChIP-seq experimental technology, the genome-wide distribution of histone methylation (such as H3K4me3, H3K27me3, H3K9me3) in M. oryzae and other filamentous fungi can be identified. Therefore, this method can help elucidate the molecular mechanisms underlying how epigenetic modifications regulate their candidate target genes during fungal pathogenesis in plant pathology.

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>acidic casein hydrolysate</td>
			<td>WAKO</td>
			<td>65072-00-6</td>
			<td>Medium configuration</td>
		</tr>
		<tr>
			<td>agar powder</td>
			<td>scientan</td>
			<td>9002-18-0</td>
			<td>Medium configuration</td>
		</tr>
		<tr>
			<td>deoxycholic acid</td>
			<td>MedChemExpress</td>
			<td>83-44-3</td>
			<td>protein and dissolution</td>
		</tr>
		<tr>
			<td>DNA End-Repair kit</td>
			<td>NovoBiotec</td>
			<td>ER81050</td>
			<td>Repair DNA or cDNA damaged by enzymatic or mechanical shearing</td>
		</tr>
		<tr>
			<td>Dynabeads</td>
			<td>Invitrogen</td>
			<td>no.100.02D</td>
			<td>Binding target</td>
		</tr>
		<tr>
			<td>EB buffer</td>
			<td>JIMEI</td>
			<td>JC2174</td>
			<td>Membrane and liquid</td>
		</tr>
		<tr>
			<td>EDTA</td>
			<td>ThermoFisher</td>
			<td>AM9912</td>
			<td>protease inhibitor</td>
		</tr>
		<tr>
			<td>enzymatic casein hydrolysate</td>
			<td>Sigma</td>
			<td>91079-40-2</td>
			<td>Medium configuration</td>
		</tr>
		<tr>
			<td>glucose</td>
			<td>Sigma</td>
			<td>50-99-7</td>
			<td>Medium configuration</td>
		</tr>
		<tr>
			<td>glycogen</td>
			<td>ThermoFisher</td>
			<td>AM9510</td>
			<td>Precipitant action</td>
		</tr>
		<tr>
			<td>H3K4me3 antibodies</td>
			<td>Abcam</td>
			<td>ab8580</td>
			<td>Immune response to H3K4me3 protein</td>
		</tr>
		<tr>
			<td>illumina Genome Analyzer</td>
			<td>illumina</td>
			<td>illumina Hiseq 2000</td>
			<td>Large configuration</td>
		</tr>
		<tr>
			<td>Illumina PCR primers</td>
			<td>illumina</td>
			<td>CleanPlex</td>
			<td>Random universal primer</td>
		</tr>
		<tr>
			<td>isoamyl alcohol</td>
			<td>chemical book</td>
			<td>30899-19-5</td>
			<td>Purified DNA</td>
		</tr>
		<tr>
			<td>LiCl</td>
			<td>ThermoFisher</td>
			<td>AM9480</td>
			<td>specific removal RNA</td>
		</tr>
		<tr>
			<td>lysing enzymes</td>
			<td>Sigma</td>
			<td>L1412-10G</td>
			<td>cell lysis buffer</td>
		</tr>
		<tr>
			<td>Mouse IgG</td>
			<td>Yeasen</td>
			<td>36111ES10</td>
			<td>Animal normal immunoglobulin</td>
		</tr>
		<tr>
			<td>NaCl solution</td>
			<td>ThermoFisher</td>
			<td>7647-14-5</td>
			<td>Medium configuration</td>
		</tr>
		<tr>
			<td>NaHCO3</td>
			<td>Seebio</td>
			<td>SH30173.08*</td>
			<td>preparation of protein complex eluent</td>
		</tr>
		<tr>
			<td>NP-40</td>
			<td>ThermoFisher</td>
			<td>85124</td>
			<td>cell lysate to promote cell lysis</td>
		</tr>
		<tr>
			<td>PCR Purification kit</td>
			<td>Qiagen</td>
			<td>28004</td>
			<td>The purification procedure removes primers from DNA samples</td>
		</tr>
		<tr>
			<td>protease inhibitors</td>
			<td>ThermoFisher</td>
			<td>A32965</td>
			<td>A protein inhibitor that decreases protein activity</td>
		</tr>
		<tr>
			<td>Proteinase K</td>
			<td>ThermoFisher</td>
			<td>AM2546</td>
			<td>DNA Extraction Reagent</td>
		</tr>
		<tr>
			<td>Qubit 4.0</td>
			<td>ThermoFisher</td>
			<td>Q33226</td>
			<td>Medium configuration</td>
		</tr>
		<tr>
			<td>RIPA buffer</td>
			<td>ThermoFisher</td>
			<td>9806S</td>
			<td>cell lysis buffer</td>
		</tr>
		<tr>
			<td>RNase A</td>
			<td>ThermoFisher</td>
			<td>AM2271</td>
			<td>Purified DNA</td>
		</tr>
		<tr>
			<td>SDS</td>
			<td>ThermoFisher</td>
			<td>AM9820</td>
			<td>cover up the charge differences</td>
		</tr>
		<tr>
			<td>sodium acetate solution</td>
			<td>ThermoFisher</td>
			<td>R1181</td>
			<td>Acetic acid buffer</td>
		</tr>
		<tr>
			<td>sodium deoxycholate</td>
			<td>ThermoFisher</td>
			<td>89904</td>
			<td>inhibition of protease degradation</td>
		</tr>
		<tr>
			<td>T4 DNA ligase</td>
			<td>ThermoFisher</td>
			<td>EL0011</td>
			<td>Under the condition of ATP as coenzyme, DNA ligase</td>
		</tr>
		<tr>
			<td>T4 DNA ligase buffer</td>
			<td>ThermoFisher</td>
			<td>B69</td>
			<td>DNA ligase buffer</td>
		</tr>
		<tr>
			<td>Tris-HCl</td>
			<td>ThermoFisher</td>
			<td>1185-53-1</td>
			<td>buffer action</td>
		</tr>
		<tr>
			<td>Triton X-100</td>
			<td>ThermoFisher</td>
			<td>HFH10</td>
			<td>keep the membrane protein stable</td>
		</tr>
		<tr>
			<td>yeast extract</td>
			<td>OXOID</td>
			<td>LP0021</td>
			<td>Medium configuration</td>
		</tr>
	</tbody>
</table>

genome-wide analysis of histone modifications distribution using the chromatin immunoprecipitation sequencing method in magnaporthe oryzae

Chromatin immunoprecipitation sequencing (ChIP-seq) is a powerful and widely used molecular technique for mapping whole genome locations of transcription factors (TFs), chromatin regulators, and histone modifications, as well as detecting entire genomes for uncovering TF binding patterns and histone posttranslational modifications. Chromatin-modifying activities, such as histone methylation, are often recruited to specific gene regulatory sequences, causing localized changes in chromatin structures and resulting in specific transcriptional effects. The rice blast is a devastating fungal disease on rice throughout the world and is a model system for studying fungus-plant interaction. However, the molecular mechanisms in how the histone modifications regulate their virulence genes in Magnaporthe oryzae remain elusive. More researchers need to use ChIP-seq to study how histone epigenetic modification regulates their target genes. ChIP-seq is also widely used to study the interaction between protein and DNA in animals and plants, but it is less used in the field of plant pathology and has not been well developed. In this paper, we describe the experimental process and operation method of ChIP-seq to identify the genome-wide distribution of histone methylation (such as H3K4me3) that binds to the functional target genes in M. oryzae. Here, we present a protocol to analyze the genome-wide distribution of histone modifications, which can identify new target genes in the pathogenesis of M. oryzae and other filamentous fungi.

The whole flow chart of the ChIP-seq method is shown in Figure 1. ChIP-seq experiments were performed using antibodies against H3K4me3 in the wild-type strain P131 and three null mutant strains that were devoid of mobre2, mospp1, and moswd2 gene to verify the whole genome-wide profile of histone H3K4me3 distribution in M. oryzae. The protoplasts of the wild-type strain, &#916;mobre2, &#916;mospp1, and &#916;moswd2, were prepared ...

Watch this Scientific Journal Video about Genome-wide Analysis of Histone Modifications Distribution using the Chromatin Immunoprecipitation Sequencing Method in Magnaporthe oryzae at JoVE.com

Genome-wide Analysis of Histone Modifications Distribution using the Chromatin Immunoprecipitation Sequencing Method in Magnaporthe oryzae

Here, we present a protocol to analyze the genome-wide distribution of histone modifications, which can identify new target genes in the pathogenesis of M. oryzae and other filamentous fungi.

Genome-wide Analysis of Histone Modifications Distribution using the Chromatin Immunoprecipitation Sequencing Method in Magnaporthe oryzae

Chromatin immunoprecipitation sequencing (ChIP-seq) is a powerful and widely used molecular technique for mapping whole genome locations of transcription ...

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Replication of the genome occurs during S phase of the cell cycle in a highly regulated process that ensures the fidelity of DNA duplication. Each genomic ...

Analysis of the c-KIT Ligand Promoter Using Chromatin Immunoprecipitation

Multiple cellular processes, including DNA replication and repair, DNA recombination, and gene expression, require interactions between proteins and DNA. ...

Chromatin Immunoprecipitation (ChIP) in Mouse T-cell Lines

Chromatin Immunoprecipitation ChIP in Mouse T-cell Lines

Signaling pathways regulate gene expression programs via the modulation of the chromatin structure at different levels, such as by post-translational ...

Generation of Native Chromatin Immunoprecipitation Sequencing Libraries for Nucleosome Density Analysis

We present a modified native chromatin immunoprecipitation sequencing (ChIP-seq) experimental protocol compatible with a Gaussian mixture distribution ...

Chromatin Immunoprecipitation (ChIP) Protocol for Low-abundance Embryonic Samples

Chromatin Immunoprecipitation ChIP Protocol for Low-abundance Embryonic Samples

Chromatin immunoprecipitation (ChIP) is a widely-used technique for mapping the localization of post-translationally modified histones, histone variants, ...

Mapping Genome-wide Accessible Chromatin in Primary Human T Lymphocytes by ATAC-Seq

Assay for Transposase-Accessible Chromatin with high-throughput sequencing (ATAC-seq) is a method used for the identification of open (accessible) regions ...

Generation of Genome-wide Chromatin Conformation Capture Libraries from Tightly Staged Early Drosophila Embryos

Generation of Genome-wide Chromatin Conformation Capture Libraries from Tightly Staged Early Drosophila Embryos

Investigating the three-dimensional architecture of chromatin offers invaluable insight into the mechanisms of gene regulation. Here, we describe a ...

Chromatin Immunoprecipitation (ChIP) of Histone Modifications from Saccharomyces cerevisiae

Chromatin Immunoprecipitation ChIP of Histone Modifications from Saccharomyces cerevisiae

Histone post-translational modifications (PTMs), such as acetylation, methylation and phosphorylation, are dynamically regulated by a series of enzymes ...

Promoter Capture Hi-C: High-resolution, Genome-wide Profiling of Promoter Interactions

The three-dimensional organization of the genome is linked to its function. For example, regulatory elements such as transcriptional enhancers control the ...

Chromatin Immunoprecipitation of Murine Brown Adipose Tissue

Most cellular processes are regulated by transcriptional modulation of specific gene programs. Such modulation is achieved through the combined actions of ...