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Tagment dna tde1 enzyme

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The Tagment DNA TDE1 Enzyme is a critical component in the Nextera DNA Library Preparation kit. It is responsible for the fragmentation and tagging of DNA samples, preparing them for subsequent library preparation steps.

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Lab products found in correlation

Ampure xp bead, by Beckman Coulter (6 mentions) Minelute pcr purification kit, by Qiagen (6 mentions) Ampure xp magnetic beads, by Beckman Coulter (2 mentions) Nextera dna flex library prep kit, by Illumina (2 mentions) Nextseq75 high output v2 kit, by Illumina (2 mentions) Nextseq 500, by Illumina (2 mentions) Nebnext high fidelity 2x pcr master mix, by New England Biolabs (2 mentions) Hiseq 3000 4000 sbs kit, by Illumina (2 mentions) E gel ex agarose gel, by Thermo Fisher Scientific (1 mentions) Hiseq kit, by Illumina (1 mentions) Hiseq 1500, by Illumina (1 mentions) High sensitivity dna bioanalyzer, by Agilent Technologies (1 mentions) Ampure xp beads, by Roche (1 mentions) Kapa hifi hotstart readymix, by Roche (1 mentions) Qubit dsdna hs kit, by Thermo Fisher Scientific (1 mentions) Novaseq 6000 machine, by Illumina (1 mentions) Agencourt ampure xp bead, by Beckman Coulter (1 mentions) Nebnext high fidelity 2 pcr master mix, by New England Biolabs (1 mentions) Td buffer, by Illumina (1 mentions) Td tagment dna buffer, by Illumina (1 mentions)

14 protocols using tagment dna tde1 enzyme

1

Omni-ATAC Profiling of MCF-7 Cells

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At the endpoint of each cell condition, we immediately performed the Omni-ATAC protocol (Corces et al., 2017 (link)) on 50,000 live MCF-7 cells, using Illumina Tagment DNA Enzyme TDE1 (20034197) at the tagmentation step and double-sided bead purification at the endpoint with Agencourt AMPure XP magnetic beads (A63880). The exact protocol we used is available in the protocols folder at https://github.com/emsanford/combined_responses_paper (Sanford, 2020a ; copy archived at swh:1:rev:e25f3d9eefd72ac1ab2885d9b0f3ad0c3cf0b3b8). We then performed paired-end sequencing using one 75-cycle NextSeq 500/550 High Output Kit v2.5 (20024906) for each replicate, yielding ~42 million read pairs per sample.
Sanford E.M., Emert B.L., Coté A, & Raj A. (2020). Gene regulation gravitates toward either addition or multiplication when combining the effects of two signals. eLife, 9, e59388.
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2

Single-cell Omni-ATAC Profiling of MCF-7 Cells

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At the endpoint of each cell condition, we immediately performed the Omni-ATAC protocol (Corces et al., 2017) (link) on 50,000 live MCF-7 cells, using Illumina Tagment DNA Enzyme TDE1 (20034197) at the tagmentation step and double-sided bead purification at the endpoint with Agencourt AMPure XP magnetic beads (A63880). The exact protocol we used is available in the protocols folder at https://github.com/emsanford/combined_responses_paper. We then performed paired-end sequencing using one 75-cycle NextSeq 500/550 High Output Kit v2.5 (20024906) for each replicate, yielding ~42 million read pairs per sample.
Sanford E.M., Emert B., Coté A., & Raj A. (2020). Gene regulation gravitates towards either addition or multiplication when combining the effects of two signals.
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3

Omni-ATAC-seq Library Preparation

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ATAC-seq was performed using the Omni-ATAC protocol64 . For each sample, 5 × 104 to 6 × 104 cells were lysed with buffer containing 0.1% NP-40, 0.1% Tween-20, and 0.01% digitonin, then the lysate was washed and incubated with Tagment DNA TDE1 Enzyme (Illumina) for 30 min at 37°C. DNA was purified with a Qiagen MinElute PCR Purification Kit. Library fragments were amplified using NEBNext® High-Fidelity 2X PCR Master Mix (NEB, M0541) and custom primers (Supplementary Table 7b) with unique dual indexes. PCR amplification cycles were in accordance with the manufacturer’s instructions. PCR products were purified using AMPure XP beads (Beckman Coulter, A63881) at a 1:1 ratio according to the manufacturer’s instructions. Purified PCR products sized from 150 bp to 1 kb were purified from 2% E-Gel EX agarose gels (Thermo Fisher, G401002). Libraries were pooled and sequenced in paired-end mode by using an Illumina HiSeq kit. Raw reads were trimmed to remove the Tn5 adaptor sequence by using skewer (version 0.2.2) then mapped to hg19 by using BWA-MEM (version 0.7.16a). Duplicated multi-mapped reads were removed with SAMtools (version 0.17). ATAC-seq peaks were called using MACS2 (version 2.1.1) with the following parameters: macs2 callpeak –nomodel –shift –100 –extsize 200. BigWiggle files were generated using DeepTools (version 3.2.0).
Feng R., Mayuranathan T., Huang P., Doerfler P.A., Li Y., Yao Y., Zhang J., Palmer L.E., Mayberry K., Christakopoulos G.E., Xu P., Li C., Cheng Y., Blobel G.A., Simon M.C, & Weiss M.J. (2022). Activation of γ-globin expression by hypoxia-inducible factor 1α. Nature, 610(7933), 783-790.
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4

OmniATAC Transposition and Sequencing

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The OmniATAC51 (link) transposition reaction was performed with 50 thousand ES cells using the Tagment DNA TDE1 Enzyme (Illumina, #20034197). DNA was purified using the PCR clean-up MinElute kit (Qiagen, #28006). The transposed DNA was subsequently amplified in 50 µl reactions with custom primers52 (link) listed in Supplementary Table 8. Libraries were purified and size selected for fragments <600 bp using the Agencourt AMPure XP beads (Beckman Coulter, #A63882). Sequencing was performed by LAFUGA on the Illumina Hiseq 1500 with 50 bp single-end reads.
Groh S., Milton A.V., Marinelli L.K., Sickinger C.V., Russo A., Bollig H., de Almeida G.P., Schmidt A., Forné I., Imhof A, & Schotta G. (2021). Morc3 silences endogenous retroviruses by enabling Daxx-mediated histone H3.3 incorporation. Nature Communications, 12, 5996.
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5

Tagmentation and Library Preparation

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Tagmentation was performed by pelleting sorted cells and resuspending in 2x Tagmentation Buffer (Illumina, #20034198), 0.02% Digitonin, and 0.1% Tween-20, with Tagment DNA TDE1 Enzyme (Illumina) and incubating at 37°C for one hour. The mix was then incubated with proteinase K for 30 minutes at 40°C.
High molecular weight DNA was removed with a 0.7x selection ratio of Agencourt AMPureXP beads (Beckman Coulter, #A63881) and low molecular weight DNA was isolated from the remainder with a 1.2x selection ratio.
Library amplification was performed with KAPA HiFi HotStart ReadyMix (KAPA, #KK2601) and i5 and i7 indexing primers (Nextera DNA CD Indexes, #20018708) for 11–15 total cycles depending on the sample concentration reached after 5 cycles (KAPA Library Quantification kit, #KK4835).
Library DNA was purified with a 1x selection ratio of AMPureXP beads. The final library was verified by High Sensitivity DNA Bioanalyzer (Agilent, #5067–4626) to ensure the presence of appropriately sized DNA fragments.
Rahmberg A.R., Markowitz T.E., Mudd J.C., Hirsch V, & Brenchley J.M. (2022). Epigenetic reprogramming leads to down regulation of CD4 and functional changes in African green monkey memory CD4+ T cells. Journal of immunology (Baltimore, Md. : 1950), 209(2), 337-345.
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6

ATAC-seq Protocol for Chromatin Accessibility

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Assay for transposase-accessible chromatin sequencing (ATACseq) was performed by the Center for Medical Genomics, Indiana University School of Medicine (Indianapolis, IN). The Tagment DNA TDE1 enzyme and Nextera DNA Flex Library Prep Kit (Illumina, catalog no., 15027866 and 15027865) were used. Briefly, 1 Â 10 5 OVCAR3 and SKOV3 cells were lysed in a nonionic detergent to yield pure nuclei. The chromatin was fragmented and simultaneously tagmented with the sequencing adaptor using Tn5 transposase to generate ATAC-seq libraries, which were sequenced on NextSeq 500 (Illumina) with NextSeq75 High Output v2 Kit (Illumina, catalog no. FC-404-2005). Raw fastq files were aligned to the human GRCH38 genome by using bowtie 2 (39) . MACS2 and ENCODE standardized pipeline and parameters were utilized for peak detection (40) . Peaks on the promoter region of the UCHL1 gene were plotted using the UCSC genome browser (41) .
Tangri A., Lighty K., Loganathan J., Mesmar F., Podicheti R., Zhang C., Iwanicki M., Drapkin R., Nakshatri H, & Mitra S. (2021). Deubiquitinase UCHL1 Maintains Protein Homeostasis through the PSMA7-APEH-Proteasome Axis in High-grade Serous Ovarian Carcinoma. Molecular cancer research : MCR, 19(7).
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7

ATAC-seq Analysis of OVCAR3 and SKOV3 Cells

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ATAC-seq was performed by the Center for Medical Genomics, Indiana University School of Medicine. The Tagment DNA TDE1 enzyme and Nextera DNA Flex Library Prep kit (Illumina cat#15027866 and 15027865) were used. Briefly, 1x10 5 OVCAR3 and SKOV3 cells were lysed in a non-ionic detergent to yield pure nuclei. The chromatin was fragmented and simultaneously tagmented with the sequencing adaptor using Tn5 transposase to generate ATAC-seq libraries, which were sequenced on NextSeq 500 (Illumina) with NextSeq75 High Output v2 kit (Illumina, cat# FC-404-2005). Raw fastq files were aligned to the human GRCH38 genome by using bowtie 2 [39] . MACS2 and ENCODE standardized pipeline and parameters were utilized for peak detection [40] . Peaks on the promoter region of the UCHL1 gene were plotted using the UCSC genome browser [41] .
Tangri A., Lighty K., Loganathan J., Mesmar F., Podicheti R., Zhang C., Iwanicki M.P., Zhang C., & Mitra S. (2020). Deubiquitinase UCHL1 Maintains Protein Homeostasis through PSMA7-APEH-Proteasome Axis in High-Grade Serous Ovarian Carcinoma.
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8

ATAC-seq Profiling of LPS and Dexamethasone Responses

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For ATAC-seq, 50.000 BMDMs were treated either with 100ng/ml LPS or PBS, and 1µM Dexamethasone or 0.1% ethanol for 3 hours. Transposition was performed using the OmniATAC protocol (Corces et al. 2017 ) and the tagment DNA TDE1 enzyme (Illumina, 20034197) . DNA was purified using the MinElute PCR purification kit (Qiagen). Afterwards, the transposed DNA was amplified using custom primers as previously described (Buenrostro et al. 2013) . Libraries were purified using the MinElute PCR purification kit (Qiagen) and size selected for fragments 150bp-600bp using the Agencourt AMPure XP beads (Beckman Coulter). The quality of the libraries was determined by the Qubit dsDNA HS kit (Thermo Scientific) and the Agilent High Sensitivity DNA 2100 Bioanalyzer. The samples were sequenced on an Illumina Novaseq 6000 machine.
Mechtidou A., Greulich F., Strickland B.A., Jouffe C., Cernilogar F.M., Schotta G., & Uhlenhaut N.H. (2021). BRG1 defines a genomic subset of inflammatory genes transcriptionally controlled by the glucocorticoid receptor.
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9

ATAC-seq for Chromatin Profiling

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Profiling of chromatin was performed by assay for transposase-accessible chromatin (ATAC)–seq as described by Buenrostro et al. (2015) (link). Briefly, 8–50,000 viably frozen T cells were thawed, washed in cold PBS, and lysed. The transposition reaction containing TDE1 Tagment DNA Enzyme (catalog # 20034198; Illumina) was incubated at 37°C for 30 min. The DNA was cleaned with the MinElute PCR Purification Kit (catalog # 28004; Qiagen), and the material was amplified for five cycles using NEBNext High-Fidelity 2× PCR Master Mix (catalog #M0541L; New England Biolabs). After evaluation by real-time PCR, 7–13 additional PCR cycles were done. The final product was cleaned by aMPure XP beads (catalog # A63882; Beckman Coulter) at a 1× ratio, and size selection was performed at a 0.5× ratio. Libraries were sequenced on a HiSeq4000 in a PE50 run, using the HiSeq 3000/4000 SBS Kit (Illumina). An average of 57 million paired reads were generated per sample.
Schad S.E., Chow A., Mangarin L., Pan H., Zhang J., Ceglia N., Caushi J.X., Malandro N., Zappasodi R., Gigoux M., Hirschhorn D., Budhu S., Amisaki M., Arniella M., Redmond D., Chaft J., Forde P.M., Gainor J.F., Hellmann M.D., Balachandran V., Shah S., Smith K.N., Pardoll D., Elemento O., Wolchok J.D, & Merghoub T. (2022). Tumor-induced double positive T cells display distinct lineage commitment mechanisms and functions. The Journal of Experimental Medicine, 219(6), e20212169.
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10

Scalable Omni-ATAC Sequencing Protocol

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The Omni-ATAC protocol was performed with 50,000 cells (or 25,000 cells for day 50 neurons) with minor modifications from (Corces et al, 2017 (link)). Using TDE1 Tagment DNA Enzyme and TD Buffer from Illumina. Cells were either derived from FACS or directly from cell culture plates after detachment with Accutase. After amplification with primer sets as described in (Buenrostro et al, 2013 (link)), libraries underwent size selection using AMPure XP beads (Beckman Coulter) to remove large fragments. Libraries were stored at −20 °C. For library quality control, the Agilent High Sensitivity DNA kit was used in a 2100 Bioanalyzer instrument. Libraries used for sequencing presented a fragment distribution starting from ~200 bp until around 1000 bp, with nucleosomal pattern peaks.
Gomez Ramos B., Ohnmacht J., de Lange N., Valceschini E., Ginolhac A., Catillon M., Ferrante D., Rakovic A., Halder R., Massart F., Arena G., Antony P., Bolognin S., Klein C., Krause R., Schulz M.H., Sauter T., Krüger R, & Sinkkonen L. (2023). Multiomics analysis identifies novel facilitators of human dopaminergic neuron differentiation. EMBO Reports, 25(1), 254-285.
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