Ampure xp beads

Manufactured by New England Biolabs

Sourced in United States

AMPure XP beads are paramagnetic beads used for the purification and size-selection of nucleic acids. They provide a simple, bead-based method for cleaning up PCR and sequencing reactions by selectively binding DNA fragments based on their size.

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Lab products found in correlation

Ampure xp bead, by Beckman Coulter (7 mentions) Blunt ta ligase master mix, by New England Biolabs (4 mentions) Hiseq 2500, by Illumina (3 mentions) Truseq stranded mrna sample preparation kit, by Illumina (3 mentions) Nextseq 550, by Illumina (2 mentions) Minelute pcr purification column, by Qiagen (2 mentions) Agilent 2100 bioanalyzer high sensitivity dna chip, by Agilent Technologies (2 mentions) 2100 bioanalyzer, by Agilent Technologies (2 mentions) Nextseq, by Illumina (2 mentions) Pfu ultra 2 fusion dna polymerase, by Agilent Technologies (2 mentions) End repair mix, by New England Biolabs (2 mentions) T4 dna ligase, by New England Biolabs (2 mentions) Ultra 2 end repair module, by New England Biolabs (1 mentions) Epitect bisulfite conversion kit, by Qiagen (1 mentions) Nuclease free water, by Merck Group (1 mentions) Qubit dsdna hs assay kit, by Thermo Fisher Scientific (1 mentions) Blunt ta ligation master mix, by New England Biolabs (1 mentions) Nebnext ultra 2 end repair da tailing kit, by New England Biolabs (1 mentions) Rneasy micro kit, by Qiagen (1 mentions) Rna 6000 pico kit, by Agilent Technologies (1 mentions)

Close spelling variants of this product

AMPure beads (9 citations)

33 protocols using ampure xp beads

Isolation and Nanopore Sequencing of ecDNA

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ecDNA was isolated by Circle-Seq [25 (link)] method, which digested linear DNA with modifications. Briefly, 10 μg of M.EcoGII treated DNA was subjected to a reaction mixture containing 1 × plasmid-safe reaction buffer, 20U plasmid-safe ATP-dependent DNase (Lucigen, E3101K), 1 mM ATP, and nuclease-free water was supplemented to a final volume of 100 μl. The reaction mixture was incubated at 37 degrees for 7 days. Every 24 h, the reaction mixture was replenished by adding 20U plasmid-safe ATP-dependent DNase, 1 mM ATP, and 0.4 μl 10X plasmid-safe reaction buffer. Digested ecDNA was purified with 1.8X AMpure XP beads (Beckman Coulter).
Purified ecDNA was prepared for nanopore sequencing by ligation kit LSK-SQK108(ONT). The samples were 10 kb by Covaris G tubes, end-repaired and dA-tailed using NEBnext Ultra II end-repair module (NEB), followed by clean-up using 1.8X AMpure XP beads. Sequencing adaptors and motor proteins were ligated to end-repaired DNA fragments using blunt/TA ligase master mix (NEB), followed by clean-up using 0.4 × AMpure XP beads. 1ug adaptor-ligated samples per flow cell were loaded onto PRO-002 flowcells and run on PromethION sequencers for up to 72 h. Data were collected by MinKNOW v.1.14.

Chen W., Weng Z., Xie Z., Xie Y., Zhang C., Chen Z., Ruan F., Wang J., Sun Y., Fang Y., Guo M., Tong Y., Li Y, & Tang C. (2021). Sequencing of methylase-accessible regions in integral circular extrachromosomal DNA reveals differences in chromatin structure. Epigenetics & Chromatin, 14, 40.

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RRBS Library Preparation with Spike-ins

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Prior to library preparation, unmethylated lambda phage DNA and SssI‐methylated T7 DNA were spiked in to genomic DNA samples to control bisulfite conversion rates. For RRBS, 20 μg of DNA was digested for 18 h by MspI and separated on 2% agarose gels. Digested DNA was excised at 150–250 and 250–350 bp. Digested DNA samples were end‐repaired and A‐tailed using the NEB Ultra Illumina library preparation kit (E7370). DNA fragments were ligated to methylated adapters (NEB E7535) following the NEB Ultra protocol. After adapter removal using Ampure XP beads (Beckman Coulter), DNA was converted using the Qiagen Epitect bisulfite conversion kit following the FFT sample protocol. After conversion, library PCR amplification was performed with 10 cycles following NEB Ultra kit protocol and cleaned up using Ampure XP beads. Two libraries were generated each for 150‐ to 250‐bp and 250‐ to 350‐bp inserts.

Manzo M., Wirz J., Ambrosi C., Villaseñor R., Roschitzki B, & Baubec T. (2017). Isoform‐specific localization of DNMT3A regulates DNA methylation fidelity at bivalent CpG islands. The EMBO Journal, 36(23), 3421-3434.

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ATAC-seq Library Generation Protocol

Cited 1 time

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To generate ATAC libraries (Corces et al., 2017 (link)), 5 × 10⁴ cells were harvested, treated with digitonin and tagmented with ~2.5 μL Tn5 Transposase for 30 min. DNA was amplified up to 10 cycles and purified using MinElute PCR purification columns (Qiagen). Following, libraries were size-selected on a 2% agarose gel (150–700bp), followed by purification using AMPure XP beads (NEB). Libraries were diluted to 5–10 ng/μL and run on an Agilent 2100 Bioanalyzer High Sensitivity DNA chip. Libraries were sequenced on Illumina NextSeq550 at 40 bp paired-end reads.

Carcamo S., Nguyen C.B., Grossi E., Filipescu D., Alpsoy A., Dhiman A., Sun D., Narang S., Imig J., Martin T.C., Parsons R., Aifantis I., Tsirigos A., Aguirre-Ghiso J.A., Dykhuizen E.C., Hasson D, & Bernstein E. (2022). Altered BAF occupancy and transcription factor dynamics in PBAF-deficient melanoma. Cell reports, 39(1), 110637.

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MinION Library Preparation and Sequencing

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Five hundred ng of PCR product from each sample, quantified with the Qubit dsDNA HS Assay Kit (Thermo Fisher Scientific), was used as the input for MinION library preparation with the Ligation Sequencing Kit 1D SQK-LSK109. DNA was end-repaired with NEBNext Ultra II End Repair/dA Tailing kit (New England Biolabs) and purified with AMPure XP beads (Beckman Coulter) in a 1:1 ratio before being eluted in 25 μl of nuclease-free water (Sigma-Aldrich). 350 ng of each cleaned DNA sample was barcoded using the Native Barcoding Expansion 1-12 and 13-24 kits (ONT: EXP-NBD104 and EXP-NBD114) and Blunt/TA Ligation Master Mix (New England Biolabs) before being purified with AMPure XP beads in a 1:1 ratio and eluted in 26 μl of nuclease-free water. The barcoded samples were pooled in equal quantities to a total of 250 ng in 45 μl of water. Sequencing adapters (AMII – ONT) were ligated to the DNA using Blunt/TA Ligation Master Mix before being purified with AMPure XP beads in a 1:2 ratio and washed with Short Fragment Buffer (SFP) (ONT) before being eluted in 15 μl of elution buffer. Samples were sequenced in two libraries on separate R9.4.1 MinION flow cells (ONT) (Table 1) without live basecalling.

Gallagher M.D., Karlsen M., Petterson E., Haugland Ø., Matejusova I, & Macqueen D.J. (2020). Genome Sequencing of SAV3 Reveals Repeated Seeding Events of Viral Strains in Norwegian Aquaculture. Frontiers in Microbiology, 11, 740.

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RNA-seq Library Preparation and Sequencing

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Cells were sorted into Trizol-LS. Total RNA was prepared using an RNeasy micro kit (Qiagen). DNase treatment was performed either on-column (Qiagen) or using a DNA-free DNA Removal kit (Life Technologies). Total RNA was quantified with RNA HS kit (Q32852) for Qubit fluorometer (Life Technologies) and analyzed for integrity using an RNA 6000 Pico kit (5067–1513) for 2100 Bioanalyzer (Agilent). mRNA was isolated from total RNA using NEBNext Poly(A) mRNA Magnetic Isolation Module (E7490; NEB). PolyA-selected mRNA was fragmented to a mean size of 300 nt, reverse transcribed to generate double-stranded cDNA, and converted to a paired-end library using NEBNext Ultra RNA Library Prep kit for Illumina (E7530; NEB) according to the manufacturer’s instructions, including the optional double size selection procedure using Agencourt AMPure XP beads. Prepared libraries were quantified with a dsDNA HS kit (Q32851) for Qubit, and the size distribution was assessed using a High Sensitivity DNA kit (5067-462) for Bioanalyzer. Libraries were sequenced on an Illumina HiSeq2500 in paired-end mode with the read length of 75 nt.

Tober J., Maijenburg M.M., Li Y., Gao L., Hadland B.K., Gao P., Minoura K., Bernstein I.D., Tan K, & Speck N.A. (2018). Maturation of hematopoietic stem cells from prehematopoietic stem cells is accompanied by up-regulation of PD-L1. The Journal of Experimental Medicine, 215(2), 645-659.

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ATAC-seq Library Generation Protocol

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Whole Genome Amplification for Sequencing

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Using the other half of the DNA sample, the steps of DNA fragmentation, blocking and nick conversion are the same as described above in NT-method. Nick-converted DNA (100 ng) was denatured by heating at 95°C for 3 min in 20 μl of H₂O, followed by adding A poly(T) tail to the ssDNA in a 30 μl reaction system containing 3 μl DNA SMART buffer (Clontech), 1 μl terminal deoxynucleotidyl transferase (TdT, Clontech) and 1 μl DNA SMART T-Tailing Mix (Clontech) by incubating at 37°C for 20 min and terminating the reaction at 70°C for 10 min. The primer annealing and template switching reaction was then performed with the Clontech SMART ChIp-seq kit (Clontech) by following the manufacturer's protocol. The final step of PCR was perfomed using the Illumina primers provided in ChIp-seq kit and 12 cycles were used for amplification. The PCR product of each sample, with unique sequencing barcode, was combined with its corresponding negative control and then size selected using AMPure XP beads (NEB). The purified library was submitted to Illumina NextSeq 500 instrument for 75 bp paired-end sequencing.

Cao B., Wu X., Zhou J., Wu H., Liu L., Zhang Q., DeMott M.S., Gu C., Wang L., You D, & Dedon P.C. (2020). Nick-seq for single-nucleotide resolution genomic maps of DNA modifications and damage. Nucleic Acids Research, 48(12), 6715-6725.

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Profiling Chromatin Accessibility in T Cell Subsets

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After FACS isolation of CD45RA⁺CD4⁺, CD45RA⁻CD4⁺, CD45RA⁺CD8⁺, and CD45RA⁻CD8⁺ T cell populations, the Fast-ATAC protocol was then performed as previously described(Corces et al., 2016 (link)). Briefly, fifty microliters of transposase mixture (25 μl of 2× TD buffer, 2.5 μl of TDE1, 0.5 μl of 1% digitonin, and 22 μl of nuclease-free water) (FC-121-1030, Illumina; G9441, Promega) was added to a cell pellet consisting of 10000-50000 cells and incubated at 37°C for 30 minutes. Transposed DNA was purified using a MinElute Reaction Cleanup kit (Qiagen), and purified DNA was eluted in 10 μl of elution buffer (10 mM Tris-HCl, pH 8). Libraries were barcoded (Nextera Index Kit, Illumina), amplified with NEBNext High Fidelity PCR Mix (New England Biolabs), and cleaned using a 1x volume of AMPure XP beads. Libraries were quantified using Agilent BioAnalyzer and sequenced on the HiSeq High Output and NovaSeq Illumina Sequencers (25 bp, paired-end).

Bachireddy P., Azizi E., Burdziak C., Nguyen V.N., Ennis C.S., Maurer K., Park C.Y., Choo Z.N., Li S., Gohil S.H., Ruthen N.G., Ge Z., Keskin D.B., Cieri N., Livak K.J., Kim H.T., Neuberg D.S., Soiffer R.J., Ritz J., Alyea E.P., Pe'er D, & Wu C.J. (2021). Mapping the evolution of T cell states during response and resistance to adoptive cellular therapy. Cell reports, 37(6), 109992.

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Oxidative Bisulfite Sequencing of mESC DNA

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mESC genomic DNA (200, 50, 10 or 1 ng) was spiked with 0.05% 4kb control methylated in CCGG sequence context and oxidised by mTet1CD as described above. Subsequently, oxidized DNA samples in 35 μl of water were reduced in a 50 μl reaction containing 600 mM sodium acetate solution (pH 4.3) and 1 M pyridine borane for 16 h at 37 °C and 850 r.p.m. in an Eppendorf ThermoMixer. The product was purified using Zymo-Spin columns. Converted samples were digested in a 20 μl reaction containing 2 U of USER enzyme (New England Biolabs) in CutSmart buffer for 1 h at 37°C and size-selected on 0.35×–1× Ampure XP beads. End-repair and A-tailing reactions, and ligation of Illumina Multiplexing adapters were prepared with KAPA Hyper kit according to the manufacturer's protocol. To prepare the control library, 200 ng of unconverted mESC gDNA with spike-in controls was digested by USER enzyme, size-selected and used for library construction as described above. The final sequencing libraries were amplified with KAPA HiFi HotStart ReadyMix for 6 cycles (for 200 ng input), 8 cycles (50 ng input), 10 cycles (10 ng input) or 14 cycles (1 ng input) and size-selected on 0.35×-1× Ampure XP beads. Final libraries were paired-end 80 bp sequenced on a NextSeq 500 sequencer (Illumina) together with other sequencing libraries.

Cheng J., Siejka-Zielińska P., Liu Y., Chandran A., Kriaucionis S, & Song C.X. (2021). Endonuclease enrichment TAPS for cost-effective genome-wide base-resolution DNA methylation detection. Nucleic Acids Research, 49(13), e76.

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Nanopore Sequencing Library Preparation

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The Nanopore SQK-LSK110 kit and EXP-PBC096 kit (Oxford Nanopore Technologies [ONT], Oxford, UK) were used to prepare the libraries. Briefly, 50 ng of each PCR product was diluted to a total volume of 24 μL for barcoding. Each reaction mix included 1 μL of ONT barcode mix, 24 μL of PCR product, and 25 μL of LongAmp Tag 2× master mix (New England Biolabs, Ipswich, MA, USA). The thermal conditions for barcoding were programmed according to the SQK-LSK110 kit instructions. The barcoded samples were further cleaned up using AMPure XP beads (Beckman Coulter, Brea, CA, USA). In each sequencing run, equal molar amounts of 16 barcoded PCR products were pooled together to a total of 1 μg. DNA repair and end-prep were done using the NEBNext FFPE DNA repair mix and the Ultra II end-prep module (New England Biolabs) according to the SQK-LSK110 kit protocol. The NEBNext Quick T4 DNA ligase module (New England Biolabs) was used for library ligation, and the short fragment buffer (SFB) was selected for washing the library during ligation cleanup with AMPure XP beads. Then 40 fmol (34.6 ng of length 1,400 bp) of DNA library was loaded to a MinION R9.4 flow cell for sequencing following the priming and loading protocol. At the end of each run, the flow cell was washed using an EXP-WSH004 kit according to the manufacturer’s protocol. The flow cell was then stored at 4°C until the next sequencing.

Li X., Tseng H.T., Hemmings G., Omolehin O., Taylor C., Taylor A., Kong P., Daughtrey M., Gouker F, & Hong C. (2023). Characterization of Boxwood Shoot Bacterial Communities and Potential Impact from Fungicide Treatments. Microbiology Spectrum, 11(2), e04163-22.

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