Name: Cattura delle rotture di siti fragili comuni mediante γH2A nativo. X ChIP (Video) | JoVE Journal
Uploaded: 2025-01-24T14:20:00.000+00:00
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Description: Scopri un metodo rapido per rilevare le rotture dei siti fragili comuni mediante ChIP nativo γH2A.X. Questa tecnica riduce tempi e lavoro mantenendo alta affidabilità nella ricerca sul cancro.

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<li>Nelson, J. D., Denisenko, O., Bomsztyk, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Fast+chromatin+immunoprecipitation+assay%5BTitle%5D)+AND+%22Nuc+Acids+Res%22%5BJournal%5D)">Fast chromatin immunoprecipitation assay.</a> Nuc Acids Res. 34 (5), e2(2006).</li>
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<li>Hebbes, T. R., Thorne, A. W., Clayton, A. L., Crane-Robinson, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Histone+acetylation+and+globin+gene+switching%5BTitle%5D)+AND+%22Nuc+Acids+Res%22%5BJournal%5D)">Histone acetylation and globin gene switching.</a> Nuc Acids Res. 20 (5), 1017-1022 (1992).</li>
<li>Hebbes, T. R., Clayton, A. L., Thorne, A. W., Crane-Robinson, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Core+histone+hyperacetylation+co-maps+with+generalized+DNase+I+sensitivity+in+the+chicken+beta-globin+chromosomal+domain%5BTitle%5D)+AND+%22EMBO+J%22%5BJournal%5D)">Core histone hyperacetylation co-maps with generalized DNase I sensitivity in the chicken beta-globin chromosomal domain.</a> EMBO J. 13 (8), 1823-1830 (1994).</li>
<li>Orlando, V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Mapping+chromosomal+proteins+in+vivo+by+formaldehyde-crosslinked-chromatin+immunoprecipitation%5BTitle%5D)+AND+%22Trend+Biochem%2C+Sci%22%5BJournal%5D)">Mapping chromosomal proteins in vivo by formaldehyde-crosslinked-chromatin immunoprecipitation.</a> Trend Biochem, Sci. 25 (3), 99-104 (2000).</li>
<li>Myers, F. A., Evans, D. R., Clayton, A. L., Thorne, A. W., Crane-Robinson, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Targeted+and+extended+acetylation+of+histones+H4+and+H3+at+active+and+inactive+genes+in+chicken+embryo+erythrocytes%5BTitle%5D)+AND+%22J+Biol+Chem%22%5BJournal%5D)">Targeted and extended acetylation of histones H4 and H3 at active and inactive genes in chicken embryo erythrocytes.</a> J Biol Chem. 276 (23), 20197-20205 (2001).</li>
<li>Litt, M. D., Simpson, M., Recillas-Targa, F., Prioleau, M. N., Felsenfeld, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Transitions+in+histone+acetylation+reveal+boundaries+of+three+separately+regulated+neighboring+loci%5BTitle%5D)+AND+%22EMBO+J%22%5BJournal%5D)">Transitions in histone acetylation reveal boundaries of three separately regulated neighboring loci.</a> EMBO J. 20 (9), 2224-2235 (2001).</li>
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<li>Clayton, A. L., Hebbes, T. R., Thorne, A. W., Crane-Robinson, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Histone+acetylation+and+gene+induction+in+human+cells%5BTitle%5D)+AND+%22FEBS+Lett%22%5BJournal%5D)">Histone acetylation and gene induction in human cells.</a> FEBS Lett. 336 (1), 23-26 (1993).</li>
<li>Tseng, Z., Wu, T., Liu, Y., Zhong, M., Xiao, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Using+native+chromatin+immunoprecipitation+to+interrogate+histone+variant+protein+deposition+in+embryonic+stem+cells%5BTitle%5D)+AND+%22Methods+Mol+Biol%22%5BJournal%5D)">Using native chromatin immunoprecipitation to interrogate histone variant protein deposition in embryonic stem cells.</a> Methods Mol Biol. 1176, 11-22 (2014).</li>
<li>Durkin, S. G., Glover, T. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Chromosome+fragile+sites%5BTitle%5D)+AND+%22Ann+Rev+Genetics%22%5BJournal%5D)">Chromosome fragile sites.</a> Ann Rev Genetics. 41, 169-192 (2007).</li>
<li>Lee, J. B., Keung, A. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Chromatin+immunoprecipitation+in+human+and+yeast+cells%5BTitle%5D)+AND+%22Methods+Mol+Biol%22%5BJournal%5D)">Chromatin immunoprecipitation in human and yeast cells.</a> Methods Mol Biol. 1767, 257-269 (2018).</li>
<li>Miyamoto, R., Yokoyama, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Protocol+for+fractionation-assisted+native+ChIP+%28fanChIP%29+to+capture+protein-protein%2FDNA+interactions+on+chromatin%5BTitle%5D)+AND+%22STAR+Protoc%22%5BJournal%5D)">Protocol for fractionation-assisted native ChIP (fanChIP) to capture protein-protein/DNA interactions on chromatin.</a> STAR Protoc. 2 (2), 100404(2021).</li>
<li>Mendez, J., Stillman, B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Chromatin+association+of+human+origin+recognition+complex%2C+cdc6%2C+and+minichromosome+maintenance+proteins+during+the+cell+cycle%3A+Assembly+of+prereplication+complexes+in+late+mitosis%5BTitle%5D)+AND+%22Mol+Cell+Biol%22%5BJournal%5D)">Chromatin association of human origin recognition complex, cdc6, and minichromosome maintenance proteins during the cell cycle: Assembly of prereplication complexes in late mitosis.</a> Mol Cell Biol. 20 (22), 8602-8612 (2000).</li>
<li>Nowak, D. E., Tian, B., Brasier, A. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Two-step+cross-linking+method+for+identification+of+NF-%CE%BAB+gene+network+by+chromatin+immunoprecipitation%5BTitle%5D)+AND+%22BioTechniques%22%5BJournal%5D)">Two-step cross-linking method for identification of NF-κB gene network by chromatin immunoprecipitation.</a> BioTechniques. 39 (5), 715-725 (2005).</li>
<li>Schmittgen, T. D., Livak, K. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Analyzing+real-time+PCR+data+by+the+comparative+C%28T%29+method%5BTitle%5D)+AND+%22Nat+Protoc%22%5BJournal%5D)">Analyzing real-time PCR data by the comparative C(T) method.</a> Nat Protoc. 3 (6), 1101-1108 (2008).</li>
<li>Lu, X., Parvathaneni, S., Hara, T., Lal, A., Sharma, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Replication+stress+induces+specific+enrichment+of+RECQ1+at+common+fragile+sites+FRA3B+and+FRA16D%5BTitle%5D)+AND+%22Mol+Cancer%22%5BJournal%5D)">Replication stress induces specific enrichment of RECQ1 at common fragile sites FRA3B and FRA16D.</a> Mol Cancer. 12, 29(2013).</li>
<li>Ruijter, J. M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Amplification+efficiency%3A+linking+baseline+and+bias+in+the+analysis+of+quantitative+PCR+data%5BTitle%5D)+AND+%22Nucl+Acids+Res%22%5BJournal%5D)">Amplification efficiency: linking baseline and bias in the analysis of quantitative PCR data.</a> Nucl Acids Res. 37 (6), e45(2009).</li>
<li>Lyu, X., Chastain, M., Chai, W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Genome-wide+mapping+and+profiling+of+%CE%B3H2AX+binding+hotspots+in+response+to+different+replication+stress+inducers%5BTitle%5D)+AND+%22BMC+Genomics%22%5BJournal%5D)">Genome-wide mapping and profiling of γH2AX binding hotspots in response to different replication stress inducers.</a> BMC Genomics. 20, 579(2019).</li>
<li>Farmer, P. B., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Molecular+epidemiology+studies+of+carcinogenic+environmental+pollutants.+Effects+of+polycyclic+aromatic+hydrocarbons+%28PAHs%29+in+environmental+pollution+on+exogenous+and+oxidative+DNA+damage%5BTitle%5D)+AND+%22Mutat+Res%22%5BJournal%5D)">Molecular epidemiology studies of carcinogenic environmental pollutants. Effects of polycyclic aromatic hydrocarbons (PAHs) in environmental pollution on exogenous and oxidative DNA damage.</a> Mutat Res. 544 (2-3), 397-402 (2003).</li>
<li>Phillips, D. H., Arlt, V. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Genotoxicity%3A+damage+to+DNA+and+its+consequences%5BTitle%5D)+AND+%22EXS%22%5BJournal%5D)">Genotoxicity: damage to DNA and its consequences.</a> EXS. 99, 87-110 (2009).</li>
<li>Nitsch, S., Schneider, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Native+ChIP%3A+Studying+the+genome-wide+distribution+of+histone+modifications+in+cells+and+tissue%5BTitle%5D)+AND+%22Meth+Mol+Biol%22%5BJournal%5D)">Native ChIP: Studying the genome-wide distribution of histone modifications in cells and tissue.</a> Meth Mol Biol. 2846, (2024).</li>
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</ol>

<table><tbody><tr><td>0,2 &micro; M membrana in nitrocellulosa</td><td>Amersham</td><td>10600011</td><td></td></tr><tr><td>Actina B</td><td>ProteinTech</td><td>20536-1-AP</td><td></td></tr><tr><td>Afidicolina</td><td>MedChemExpress</td><td>HY-N6733</td><td></td></tr><tr><td>Perle magnetiche A / G di proteina di grado ChIP</td><td>ThermoFisher</td><td>26162</td><td></td></tr><tr><td>Substrato ECL occidentale Clarity</td><td>Bio-Rad</td><td>#1705061</td><td></td></tr><tr><td>Glicogeno di grado di biologia molecolare</td><td>ThermoFisher</td><td>Gatto. No. R0561</td><td></td></tr><tr><td>Anticorpo secondario coniugato HRP&nbsp;</td><td>ProteinTech</td><td>SA00001-2</td><td></td></tr><tr><td>idrossiurea&nbsp;</td><td>MedChemExpress</td><td>HY-B0313</td><td></td></tr><tr><td>Nucleasi micrococcica</td><td>NEB</td><td>Codice: M0247S</td><td></td></tr><tr><td>IgG normali</td><td>Santa Cruz</td><td>SC-2025</td><td></td></tr><tr><td>Taq Universale SYBR Verde Supermix&nbsp;</td><td>BioRad</td><td>1725120</td><td></td></tr><tr><td>&gamma; H2A. Anticorpo X&nbsp; (per ChIP)</td><td>Sigma-Aldrich</td><td>05-636</td><td></td></tr><tr><td>&gamma; H2A. Anticorpo X (per WB)</td><td>Segnalazione cellulare</td><td>#25955</td><td></td></tr></tbody></table>

cattura delle rotture di siti fragili comuni mediante γh2a nativo. x chip

Capturing Common Fragile Site Breaks by Native &#947;H2A.X ChIP

La dimensione dei frammenti di cromatina è fondamentale per il successo del ChIP nativo, in quanto influisce direttamente sull'accessibilità delle regioni del DNA per il legame degli anticorpi. Per determinare la concentrazione ottimale di MNasi per la frammentazione della cromatina, abbiamo preparato una serie di provette per microcentrifuga contenenti concentrazioni variabili di MNasi (cioè 0,0625 U, 0,125 U, 0,25 U, 0,5 U, 1 U, 2 U, 4 U, 8 U per reazione) e 40 μL di nuclei isolati. Ogni reazione è stata incubata a 37 ...

Cattura delle rotture di siti fragili comuni mediante γH2A nativo. X ChIP

Replication stress induced by exposure to extrinsic agents can lead to DNA breaks at common fragile sites, which are regions in the genome known to be ...

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Riportiamo un protocollo per combinare la metrologia atomica del microscopio a scansione a effetto tunnel per la modellazione delle superfici con la deposizione selettiva di strati atomici e l'incisione ionica reattiva. Utilizzando un robusto processo che coinvolge numerose esposizioni atmosferiche e trasporto, vengono fabbricate nanostrutture 3D con metrologia atomica.

Atomicamente tracciabile nanostrutture

Qui introduciamo protocolli sperimentali per l'osservazione in tempo reale di un processo auto-assemblaggio mediante microscopia elettronica a trasmissione di liquido-cella.

Microscopia elettronica in trasmissione di liquidi-cellula per l'inseguimento auto-assemblaggio delle nanoparticelle

Qui, presentiamo un protocollo che descrive la fabbricazione di estensibile, bicanale, dispositivi organo chip microfluidici cella cultura per ricapitolare la funzionalità a livello dell'organo in vitro.

Fabbricazione scalabile di Stretchable, Dual Channel, organo di Microfluidic chip

Questo documento descrive un protocollo insieme a uno studio comparativo di due tecniche di fabbricazione microfluidica, vale a dire la fotolitografia / incisione a umido / legame termico e l'incisione selettiva indotta dal laser (LES), che sono adatte per condizioni di alta pressione. Queste tecniche costituiscono piattaforme abilitanti per l'osservazione diretta del flusso di fluido in mezzi permeabili surrogati e sistemi fratturati in condizioni di serbatoio.

Tecniche di fabbricazione microfluidiche per test ad alta pressione del trasporto supercritico in schiuma di CO2 su microscala in serbatoi non convenzionali fratturati

Qui presentiamo un protocollo per l'esecuzione di esperimenti di reazione a gas a celle chiuse TEM in situ, dettagliando diversi metodi di preparazione del campione comunemente usati.

Esecuzione di reazioni di gas a celle chiuse in situ nel microscopio elettronico a trasmissione

Viene fornito un protocollo per la coltura e la manipolazione del tessuto embrionale di topo su un chip microfluidico. Applicando impulsi di modulatori di percorso, questo sistema può essere utilizzato per controllare esternamente le oscillazioni di segnalazione per lo studio funzionale della somitogenesi del topo.

Un approccio microfluidico per l'indagine funzionale delle oscillazioni di segnalazione che governano la somitogenesi

Questo articolo ripercorre il ciclo di vita di un risonatore a membrana in nitruro di silicio, dalla progettazione alla fabbricazione fino alla caratterizzazione.

Fabbricazione e caratterizzazione di risonatori a membrana in nitruro di silicio ad alto Q

Un nuovo protocollo di sigillatura per produrre sigilli impermeabili a base di silicone per membrane di dispositivi a chip polidimetilsilossano (PDMS). Destinato a laboratori, configurazione di modelli di chip e sviluppo di piattaforme chip su piccola scala. Dimostriamo il protocollo utilizzando membrane tissutali plastiche e native. Questa procedura richiede solo PDMS, toluene, muffa, membrana e una camera a vuoto.

Procedura accessibile di sigillatura chip-membrana in silicone per un legame flessibile e affidabile

Scopri un metodo rapido per rilevare le rotture dei siti fragili comuni mediante ChIP nativo γH2A.X. Questa tecnica riduce tempi e lavoro mantenendo alta affidabilità nella ricerca sul cancro.

Cattura delle rotture di siti fragili comuni mediante γH2A nativo. X ChIP

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