E220 evolution focused ultrasonicator

Manufactured by Covaris

Sourced in United States

The E220 evolution Focused-ultrasonicator is a lab equipment product designed for sample preparation and processing. It utilizes focused-ultrasonic energy to facilitate various applications within the scientific and research domains.

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Lab products found in correlation

Qiaquick pcr purification kit, by Qiagen (3 mentions) Cfx96 real time system, by Bio-Rad (2 mentions) Iq sybr green supermix, by Bio-Rad (2 mentions) Dynabeads protein g, by Thermo Fisher Scientific (2 mentions) Dynabead, by Thermo Fisher Scientific (2 mentions) Proteinase k, by Qiagen (2 mentions) Mgieasy fs dna library prep kit, by Covaris (2 mentions) Dna clean concentrator 5, by Zymo Research (1 mentions) Anti myc monoclonal antibody, by Takara Bio (1 mentions) Qiaamp dna mini kit, by Qiagen (1 mentions) Ht 29 cell, by American Type Culture Collection (1 mentions) Mccoy s 5a medium, by Lonza (1 mentions) Nbp1 32440, by Novus Biologicals (1 mentions) H3k27ac, by Diagenode (1 mentions)

Spelling variants of this product

Ultrasonicator (113 citations) E220 Focused-ultrasonicator (87 citations) Focused-ultrasonicator (73 citations) E220 ultrasonicator (36 citations) E220 Evolution (13 citations) E220 focused (6 citations)

8 protocols using e220 evolution focused ultrasonicator

Robust Methylation Profiling in Neuroblastoma

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Neuroblastoma cells were grown for 72 h at atmospheric conditions, and the extracted gDNA was sonicated on E220 Evolution Focused-ultrasonicator (Covaris) in 10 mM Tris-HCl (pH 8.5) buffer to yield fragments with a peak size of ∼200 bp. Five hundred nanograms of gDNA were incubated with 0.5 μM eM.SssI in 30 μl 10 mM Tris–HCl (pH 7.4), 50 mM NaCl, 0.5 mM EDTA reaction mixture, supplemented with 200 μM Ado-6-azide cofactor for 1 h at 30°C followed by thermal inactivation at 65°C for 20 min and Proteinase K treatment (0.2 mg/ml) for 30 min at 55°C, and then column purified (DNA Clean & Concentrator-5, Zymo Research, Irvine, CA, United States). After ligation of adaptors to the azide-tagged DNA, alkyne-containing DNA oligonucleotide with biotin was attached, biotinylated DNA was enriched with streptavidin-coupled magnetic beads and subsequently used in a DNA priming reaction as described previously for hmTOP-seq library preparation (Gibas et al., 2020 (link)). uTOP-seq DNA libraries were amplified for 11 cycles, size selected for ∼300 bp fragments (MagJET NGS Cleanup and Size Selection kit, TS) and subjected to Ion Proton (TS) sequencing.

Narmontė M., Gibas P., Daniūnaitė K., Gordevičius J, & Kriukienė E. (2021). Multiomics Analysis of Neuroblastoma Cells Reveals a Diversity of Malignant Transformations. Frontiers in Cell and Developmental Biology, 9, 727353.

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Epitope Tagging for Chromatin Immunoprecipitation

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Epitope tagging of proteins for ChIP experiments was carried out as previously described (Tran et al. 2017 (link)). Briefly, cells were grown in 50 ml of YEPD (YEP with dextrose) overnight and harvested at an OD₆₆₀ of 0.5. Cells were cross-linked with 1% formaldehyde for 10 min. Chromatin purification was carried out as described (Paeschke et al. 2011 (link)), except that DNA was sheared to an average size of 300 bp using an E220 evolution Focused-ultrasonicator (Covaris, Woburn, MA). Anti-MYC monoclonal antibody (#631206; Clontech) was diluted to 0.02 μg/μl and coupled to 80 μl of Dynabeads protein G (#10004D; Thermo Fisher Scientific). Reverse-cross-linking DNA was performed and purified by QIAquick PCR Purification kit (#28106; QIAGEN, Valencia, CA). Immunoprecipitated chromatin and input DNA were analyzed by quantitative PCR (qPCR) using iQ SYBR Green Supermix (#170–8882; Bio-Rad, Hercules, CA) and a CFX96 real-time system (Bio-Rad). All ChIP experiments were repeated at least three times. Strains and primers are listed in Table S1 and S2. WT cells without a Myc-tagged protein were used as a negative control. Most ChIP-qPCR were quantified by [(ChIP/Input)_{Target site}/(ChIP/Input)_YBL028C]. YBL028C is a control sequence that has very low ScPif1 and Rrm3 binding.

Chen C.F., Pohl T.J., Pott S, & Zakian V.A. (2018). Two Pif1 Family DNA Helicases Cooperate in Centromere Replication and Segregation in Saccharomyces cerevisiae. Genetics, 211(1), 105-119.

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HT-29 Cells Genomic DNA Fragmentation

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HT-29 cells (ATCC) were grown in McCoy’s 5A Medium (Lonza). Genomic DNA was isolated from the cells using the QIAamp^® DNA Mini Kit (Qiagen). The genomic DNA was sheared using an E220evolution™ focused-ultrasonicator (Covaris) to give a target fragment size of 100 bp. The resulting fragments ranged from 25 to 500 bp in length with the largest proportion of DNA fragments between 50 and 300 bp in length, as shown in Figure S1 in the Supplementary Information.

Tam B.E., Hao Y, & Sikes H.D. (2018). An Examination of Critical Parameters in Hybridization-Based Epigenotyping using Magnetic Microparticles. Biotechnology progress, 34(6), 1589-1595.

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ChIP-seq of PAX8 and H3K27ac

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Chromatin immunoprecipitation sequencing (ChIP-seq) was performed based on the methods of Schmidt et al. [36 (link)]. Cells were fixed in 1% formaldehyde for 10 minutes, and quenched with glycine. Cells were harvested, lysed in a sarkosyl-containing buffer, and sonicated using the Covaris E220evolution Focused-Ultrasonicator. 10 μg of an antibody raised against PAX8 (NBP1-32440, Novus) or 5 μg of an antibody raised against lysine 27 acetylated histone 3 (H3K27ac) (C15410196, Diagenode) was incubated with 100 μg and 4ug, respectively, of chromatin at 4°C overnight. Blocked magnetic Dynabeads (Life Technologies) were then added to the antibody-lysate conjugates and incubated at 4°C for 4 hours with rotation. Beads were then washed with RIPA buffer and treated with RNase and proteinase K (both Qiagen). DNA was eluted from the beads in Tris-EDTA buffer and cleaned up using the QIAquick PCR Purification kit (QIAgen). For each cell line two independent immunoprecipitations and one input sample were submitted for next-generation sequencing at the USC Epigenome Core Facility.

Adler E.K., Corona R.I., Lee J.M., Rodriguez-Malave N., Mhawech-Fauceglia P., Sowter H., Hazelett D.J., Lawrenson K, & Gayther S.A. (2017). The PAX8 cistrome in epithelial ovarian cancer. Oncotarget, 8(65), 108316-108332.

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Epitope Tagging for ChIP Experiments

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Epitope tagging of proteins for ChIP experiments was done as previously described7 (link)8 (link). Briefly, cells were grown overnight and harvested at an OD₆₆₀ of 0.5. Cells were crosslinked with 1% formaldehyde for 10 min. Chromatin purification was done as described7 (link)8 (link), except that DNA was sheared to an average size of 300 bps using E220 evolution Focused-ultrasonicator (Covaris, MA, USA). Anti-MYC monoclonal antibody (Clontech #631206) was diluted to 0.02 μg μl⁻¹ and coupled to 80 μl of Dynabeads protein G (ThermoFisher #10004D). After crosslinking reversal and DNA purification, the immunoprecipitated and input DNA were analysed by qPCR using iQ SYBR Green Supermix (Bio-Rad #170-8882) and CFX96 real-time system (Bio-Rad). Samples were analysed in triplicates on three independent ChIP samples for each genotype. Strains and primers are listed in Supplementary Table 9. Wild-type YPH499 cells without an Myc-tagged protein were used as a control.

Tran P.L., Pohl T.J., Chen C.F., Chan A., Pott S, & Zakian V.A. (2017). PIF1 family DNA helicases suppress R-loop mediated genome instability at tRNA genes. Nature Communications, 8, 15025.

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Optimized ChIP-seq Protocol for H3K27ac

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Our ChIP protocol was based on the methods of Schmidt et al. ^{19 (link)} Four 15cm dishes of cells were fixed in formaldehyde for 10 minutes, before quenching the fixation with glycine. Cells were harvested, lysed in a sarkosyl-containing lysis buffer, and sonicated using the Covaris E220 evolution Focused-Ultrasonicator to yield 100-300bp genomic DNA fragments. 5 μg of an antibody raised against H3K27ac (Diagenode) was incubated with blocked magnetic Dynabeads (Life Technologies) at 4°C for 4 hours. Antibody-bead conjugates were incubated with 100 μg chromatin at 4°C overnight, with constant agitation. Beads were washed extensively with RIPA buffer and then RNase and proteinase K (both Qiagen) treated. DNA was then eluted from the beads in TE buffer and cleaned up using the QIAquick PCR Purification kit (Qiagen). Two independent immunoprecipitations and one input sample were sequenced for each cell line and each sample was quality checked by site-specific qPCR prior to next generation sequencing (NGS).

Phelan C.M., Kuchenbaecker K.B., Tyrer J.P., Kar S.P., Lawrenson K., Winham S.J., Dennis J., Pirie A., Riggan M., Chornokur G., Earp M.A., Lyra PC J.r., Lee J.M., Coetzee S., Beesley J., McGuffog L., Soucy P., Dicks E., Lee A., Barrowdale D., Lecarpentier J., Leslie G., Aalfs C.M., Aben K.K., Adams M., Adlard J., Andrulis I.L., Anton-Culver H., Antonenkova N., Aravantinos G., Arnold N., Arun B.K., Arver B., Azzollini J., Balmaña J., Banerjee S.N., Barjhoux L., Barkardottir R.B., Bean Y., Beckmann M.W., Beeghly-Fadiel A., Benitez J., Bermisheva M., Bernardini M.Q., Birrer M.J., Bjorge L., Black A., Blankstein K., Blok M.J., Bodelon C., Bogdanova N., Bojesen A., Bonanni B., Borg Å., Bradbury A.R., Brenton J.D., Brewer C., Brinton L., Broberg P., Brooks-Wilson A., Bruinsma F., Brunet J., Buecher B., Butzow R., Buys S.S., Caldes T., Caligo M.A., Campbell I., Cannioto R., Carney M.E., Cescon T., Chan S.B., Chang-Claude J., Chanock S., Chen X.Q., Chiew Y.E., Chiquette J., Chung W.K., Claes K.B., Conner T., Cook L.S., Cook J., Cramer D.W., Cunningham J.M., D’Aloisio A.A., Daly M.B., Damiola F., Damirovna S.D., Dansonka-Mieszkowska A., Dao F., Davidson R., DeFazio A., Delnatte C., Doheny K.F., Diez O., Ding Y.C., Doherty J.A., Domchek S.M., Dorfling C.M., Dörk T., Dossus L., Duran M., Dürst M., Dworniczak B., Eccles D., Edwards T., Eeles R., Eilber U., Ejlertsen B., Ekici A.B., Ellis S., Elvira M., Eng K.H., Engel C., Evans D.G., Fasching P.A., Ferguson S., Ferrer S.F., Flanagan J.M., Fogarty Z.C., Fortner R.T., Fostira F., Foulkes W.D., Fountzilas G., Fridley B.L., Friebel T.M., Friedman E., Frost D., Ganz P.A., Garber J., García M.J., Garcia-Barberan V., Gehrig A., Gentry-Maharaj A., Gerdes A.M., Giles G.G., Glasspool R., Glendon G., Godwin A.K., Goldgar D.E., Goranova T., Gore M., Greene M.H., Gronwald J., Gruber S., Hahnen E., Haiman C.A., Håkansson N., Hamann U., Hansen T.V., Harrington P.A., Harris H.R., Hauke J., Hein A., Henderson A., Hildebrandt M.A., Hillemanns P., Hodgson S., Høgdall C.K., Høgdall E., Hogervorst F.B., Holland H., Hooning M.J., Hosking K., Huang R.Y., Hulick P.J., Hung J., Hunter D.J., Huntsman D.G., Huzarski T., Imyanitov E.N., Isaacs C., Iversen E.S., Izatt L., Izquierdo A., Jakubowska A., James P., Janavicius R., Jernetz M., Jensen A., Jensen U.B., John E.M., Johnatty S., Jones M.E., Kannisto P., Karlan B.Y., Karnezis A., Kast K., Kennedy C.J., Khusnutdinova E., Kiemeney L.A., Kiiski J.I., Kim S.W., Kjaer S.K., Köbel M., Kopperud R.K., Kruse T.A., Kupryjanczyk J., Kwong A., Laitman Y., Lambrechts D., Larrañaga N., Larson M.C., Lazaro C., Le N.D., Le Marchand L., Lee J.W., Lele S.B., Leminen A., Leroux D., Lester J., Lesueur F., Levine D.A., Liang D., Liebrich C., Lilyquist J., Lipworth L., Lissowska J., Lu K.H., Lubiński J., Luccarini C., Lundvall L., Mai P.L., Mendoza-Fandiño G., Manoukian S., Massuger L.F., May T., Mazoyer S., McAlpine J.N., McGuire V., McLaughlin J.R., McNeish I., Meijers-Heijboer H., Meindl A., Menon U., Mensenkamp A.R., Merritt M.A., Milne R.L., Mitchell G., Modugno F., Moes-Sosnowska J., Moffitt M., Montagna M., Moysich K.B., Mulligan A.M., Musinsky J., Nathanson K.L., Nedergaard L., Ness R.B., Neuhausen S.L., Nevanlinna H., Niederacher D., Nussbaum R.L., Odunsi K., Olah E., Olopade O.I., Olsson H., Olswold C., O’Malley D.M., Ong K.R., Onland-Moret N.C., Orr N., Orsulic S., Osorio A., Palli D., Papi L., Park-Simon T.W., Paul J., Pearce C.L., Pedersen I.S., Peeters P.H., Peissel B., Peixoto A., Pejovic T., Pelttari L.M., Permuth J.B., Peterlongo P., Pezzani L., Pfeiler G., Phillips K.A., Piedmonte M., Pike M.C., Piskorz A.M., Poblete S.R., Pocza T., Poole E.M., Poppe B., Porteous M.E., Prieur F., Prokofyeva D., Pugh E., Pujana M.A., Pujol P., Radice P., Rantala J., Rappaport-Fuerhauser C., Rennert G., Rhiem K., Rice P., Richardson A., Robson M., Rodriguez G.C., Rodríguez-Antona C., Romm J., Rookus M.A., Rossing M.A., Rothstein J.H., Rudolph A., Runnebaum I.B., Salvesen H.B., Sandler D.P., Schoemaker M.J., Senter L., Setiawan V.W., Severi G., Sharma P., Shelford T., Siddiqui N., Side L.E., Sieh W., Singer C.F., Sobol H., Song H., Southey M.C., Spurdle A.B., Stadler Z., Steinemann D., Stoppa-Lyonnet D., Sucheston-Campbell L.E., Sukiennicki G., Sutphen R., Sutter C., Swerdlow A.J., Szabo C.I., Szafron L., Tan Y.Y., Taylor J.A., Tea M.K., Teixeira M.R., Teo S.H., Terry K.L., Thompson P.J., Thomsen L.C., Thull D.L., Tihomirova L., Tinker A.V., Tischkowitz M., Tognazzo S., Toland A.E., Tone A., Trabert B., Travis R.C., Trichopoulou A., Tung N., Tworoger S.S., van Altena A.M., Van Den Berg D., van der Hout A.H., van der Luijt R.B., Van Heetvelde M., Van Nieuwenhuysen E., van Rensburg E.J., Vanderstichele A., Varon-Mateeva R., Ana V., Edwards D.V., Vergote I., Vierkant R.A., Vijai J., Vratimos A., Walker L., Walsh C., Wand D., Wang-Gohrke S., Wappenschmidt B., Webb P.M., Weinberg C.R., Weitzel J.N., Wentzensen N., Whittemore A.S., Wijnen J.T., Wilkens L.R., Wolk A., Woo M., Wu X., Wu A.H., Yang H., Yannoukakos D., Ziogas A., Zorn K.K., Narod S.A., Easton D.F., Amos C.I., Schildkraut J.M., Ramus S.J., Ottini L., Goodman M.T., Park S.K., Kelemen L.E., Risch H.A., Thomassen M., Offit K., Simard J., Schmutzler R.K., Hazelett D., Monteiro A.N., Couch F.J., Berchuck A., Chenevix-Trench G., Goode E.L., Sellers T.A., Gayther S.A., Antoniou A.C, & Pharoah P.D. (2017). Identification of twelve new susceptibility loci for different histotypes of epithelial ovarian cancer. Nature genetics, 49(5), 680-691.

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Whole Genome Sequencing Library Prep

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For small insert-size libraries, genomic DNA was sheared (300~500-bp) with the Covaris E220 Evolution Focused-Ultrasonicator (Covaris, Inc., Woburn, MA) and subjected for library construction using the MGIEasy FS DNA Library Prep kit according to the manufacturer’s protocol. Libraries were pooled and sequenced to paired-end 100-bp with a read-depth of ~4-fold for each sample on an MGISEQ-2000 platform (MGI Tech Co., Ltd., Shenzhen, China).
For mate-pair libraries, 1-μg of genomic DNA was sheared (3~8-kb) by a HydroShear device (Digilab, Inc., Hopkinton, MA)^{22 (link)}, and subjected for library construction as previously described (Supplementary methods)²⁸. Samples were pooled and sequenced on an MGISEQ-2000 platform (MGI) for a read-depth of ~4-fold (paired-end 100-bp) for each sample.

Dong Z., Chau M.H., Zhang Y., Yang Z., Shi M., Wah Y.M., Kwok Y.K., Leung T.Y., Morton C.C, & Choy K.W. (2021). Low-pass genome sequencing-based detection of absence of heterozygosity: validation in clinical cytogenetics. Genetics in medicine : official journal of the American College of Medical Genetics, 23(7), 1225-1233.

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Whole Genome Sequencing Library Prep

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