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Nebnext ultra 2 dna library prep kit

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The NEBNext Ultra II DNA Library Prep Kit is a laboratory equipment product designed for the preparation of DNA libraries for next-generation sequencing. It provides a streamlined workflow for efficient and reliable library construction.

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Lab products found in correlation

Nextseq 500, by Illumina (15 mentions) Novaseq 6000, by Illumina (13 mentions) Hiseq 4000, by Illumina (10 mentions) Nebnext multiplex oligo, by Illumina (9 mentions) Dneasy blood and tissue kit, by Qiagen (7 mentions) Hiseq 2500, by Illumina (7 mentions) Novaseq 6000 platform, by Illumina (7 mentions) Ampure xp bead, by Beckman Coulter (6 mentions) Novaseq platform, by Illumina (6 mentions) Nextseq 550, by Illumina (6 mentions) Nextseq 500 platform, by Illumina (6 mentions) Qiaamp dna mini kit, by Qiagen (5 mentions) Agilent 2100 bioanalyzer, by Agilent Technologies (5 mentions) Miseq platform, by Illumina (5 mentions) Bioruptor pico, by Diagenode (5 mentions) Nextseq sequencer, by Illumina (5 mentions) Bbcl2fastq software, by Illumina (5 mentions) Hiseq 2500 platform, by Illumina (5 mentions) Nextseq, by Illumina (5 mentions) Hiseq x, by Illumina (4 mentions)

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NEBNext Ultra Library Prep Kit (5 citations) NEBNext library prep kit (2 citations) NEBNext Ultra II (2 citations)

242 protocols using nebnext ultra 2 dna library prep kit

1

Genomic DNA Extraction and Library Preparation

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Cells were harvested by trypsinization, spun down at 1100 x g for 1 min and the supernatant was discarded. Then, cells were spun down, resuspended in 200 μl PBS and gDNA was purified using the DNeasy Blood and Tissue kit (Quiagen). DNA was transferred to Covaris microTUBE (Covaris) and sheared on a Covaris S2 instrument (Duty cycle 10 %, Intensity 5.0, Cycles/burst 200) for 25 s. Double size selection was performed by employing AMPure XP beads (Beckman Coulter) first at 0.8-fold sample volume according to the standard protocol, followed by transfer of the supernatant and bead application at 0.12-fold sample volume. DNA library preparation was performed with the resulting DNA using the NEBNext Ultra II DNA library prep kit for Illumina [NEB] according to the standard protocol. The unamplified libraries were then treated using OsO4 (see below) and amplified according the NEBNext Ultra II DNA library prep kit for Illumina [NEB]. The finished libraries were purified using AMPure XP beads (Beckman Coulter) at 0.9x sample volume following the standard protocol.
Mitter M., Gasser C., Takacs Z., Langer C.C., Tang W., Jessberger G., Beales C.T., Neuner E., Ameres S.L., Peters J.M., Goloborodko A., Micura R, & Gerlich D.W. (2020). Conformation of sister chromatids in the replicated human genome. Nature, 586(7827), 139-144.
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2

Genomic DNA Extraction and Library Preparation

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Cells were harvested by trypsinization, spun down at 1100 x g for 1 min and the supernatant was discarded. Then, cells were spun down, resuspended in 200 μl PBS and gDNA was purified using the DNeasy Blood and Tissue kit (Quiagen). DNA was transferred to Covaris microTUBE (Covaris) and sheared on a Covaris S2 instrument (Duty cycle 10 %, Intensity 5.0, Cycles/burst 200) for 25 s. Double size selection was performed by employing AMPure XP beads (Beckman Coulter) first at 0.8-fold sample volume according to the standard protocol, followed by transfer of the supernatant and bead application at 0.12-fold sample volume. DNA library preparation was performed with the resulting DNA using the NEBNext Ultra II DNA library prep kit for Illumina [NEB] according to the standard protocol. The unamplified libraries were then treated using OsO4 (see below) and amplified according the NEBNext Ultra II DNA library prep kit for Illumina [NEB]. The finished libraries were purified using AMPure XP beads (Beckman Coulter) at 0.9x sample volume following the standard protocol.
Mitter M., Gasser C., Takacs Z., Langer C.C., Tang W., Jessberger G., Beales C.T., Neuner E., Ameres S.L., Peters J.M., Goloborodko A., Micura R, & Gerlich D.W. (2020). Conformation of sister chromatids in the replicated human genome. Nature, 586(7827), 139-144.
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3

mRNA Sequencing Library Preparation

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Total RNA was isolated with TRIzol reagent. Poly(A)+ RNA enrichment was performed with Dynabeads Oligo(dT)25 (Thermo Fisher), with two consecutive purifications according to the manufacturer’s instructions. Next, cDNA was prepared using NEBNext Ultra II RNA First and Second Strand Synthesis Module. The cDNA was purified with AmpureXP beads and library was prepared with NEBNext Ultra II DNA Library Prep Kit Illumina (NEB) according to the protocol and sequenced on a HiSeq2500 (Illumina) in single-read 50 mode.
Batki J., Schnabl J., Wang J., Handler D., Andreev V.I., Stieger C.E., Novatchkova M., Lampersberger L., Kauneckaite K., Xie W., Mechtler K., Patel D.J, & Brennecke J. (2019). The nascent RNA binding complex SFiNX licenses piRNA-guided heterochromatin formation. Nature structural & molecular biology, 26(8), 720-731.
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4

mRNA Sequencing Library Preparation

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Total RNA was isolated with TRIzol reagent. Poly(A)+ RNA enrichment was performed with Dynabeads Oligo(dT)25 (Thermo Fisher), with two consecutive purifications according to the manufacturer’s instructions. Next, cDNA was prepared using NEBNext Ultra II RNA First and Second Strand Synthesis Module. The cDNA was purified with AmpureXP beads and library was prepared with NEBNext Ultra II DNA Library Prep Kit Illumina (NEB) according to the protocol and sequenced on a HiSeq2500 (Illumina) in single-read 50 mode.
Batki J., Schnabl J., Wang J., Handler D., Andreev V.I., Stieger C.E., Novatchkova M., Lampersberger L., Kauneckaite K., Xie W., Mechtler K., Patel D.J, & Brennecke J. (2019). The nascent RNA binding complex SFiNX licenses piRNA-guided heterochromatin formation. Nature structural & molecular biology, 26(8), 720-731.
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5

Shotgun Metagenomics of Cheese Samples

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A minimum of 10 μL of DNA with a concentration between 20 and 300 ng/μL, representing at least 200 ng of DNA per sample, was used for shotgun metagenomics sequencing of cheese samples from selected ripening time points. Libraries were prepared using the NEB Ultra II DNA kit following the procedures described in the NEBNext®UltraTM II DNA Library Prep Kit for Illumina® (E7645, E7103) (Illumina 2020, Illumina, San Diego, CA, USA). All samples were sequenced on Illumina HiSeq 2500 (Illumina, San Diego, CA, USA) high throughout an 8-lane flow cell for paired ends of 2 × 125 bp. Using this system together with the Rapid Run mode flow cell, a total of 150–170 million reads were generated, facilitating multiplexing samples in the same lane (reducing the total cost of the experiment per sample).
Mohamed H.M., Barzideh Z., Siddiqi M, & LaPointe G. (2023). Taxonomy, Sequence Variance and Functional Profiling of the Microbial Community of Long-Ripened Cheddar Cheese Using Shotgun Metagenomics. Microorganisms, 11(8), 2052.
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6

Genomic Characterization of MRSA Isolates

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The UltraClean Microbial Kit (Qiagen, NW, Germany) were used to extract bacterial genomic DNA. A paired-end library with an average insert size of 350 bp ranging from 150 to 600 kb was sequenced on a HiSeq sequencer (Illumina, CA, United States). The NEBNext® UltraTM II DNA Library Prep Kit for Illumina® was used to construct sequencing libraries and then loaded onto NovaSeq S4 flow cell. The WGS data were used for genotypic characterization and FA resistance determinants analysis. Multilocus sequence typing (MLST) was performed by submitting sequences to the MLST database website (https://cge.cbs.dtu.dk/services/MLST/). Spa typing was performed by using the spa database website spaTyper (https://cge.cbs.dtu.dk/services/spatyper/). Staphylococcus cassette chromosome mec (SCCmec) typing was performed by using the database website SCCmecFinder (https://cge.cbs.dtu.dk/services/SCCmecFinder/). Resistance-associated mutations in fusA or fusE, and acquired FA resistance genes, including fusB, fusC, and fusD were performed by using the database website ResFinder (https://cge.cbs.dtu.dk/services/ResFinder/). The Illumina sequences of the 74 MRSA isolates in this study are available in the Sequence Read Archive (BioProject ID: PRJNA764055).
Zhao H., Wang X., Wang B., Xu Y., Rao L., Wan B., Guo Y., Wu X., Yu J., Chen L., Li M, & Yu F. (2021). The Prevalence and Determinants of Fusidic Acid Resistance Among Methicillin-Resistant Staphylococcus aureus Clinical Isolates in China. Frontiers in Medicine, 8, 761894.
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7

CUT&RUN for Histone Modifications

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About 2500 cells were counted for CM150-post and WM266-4 from FACS-sorted cells for each antibody (IgG, HEK27Me3 and H3K27Ac), with 500,000 cells counted for the baseline. CUT&RUN experiments were conducted following the manufacturer’s protocol with slight modifications (Epicypher, Durham, NC, USA) [44 ]. Eluted DNA libraries were prepared using the NEBNext UltraTM II DNA Library Prep Kit for Illumina (NEB #E7645S/L). Replicated sequencing libraries (n = 2) were sent to Genohub (Austin, TX, USA) for sequencing. A total of 35 uniquely barcoded libraries from the CUT and RUN protocol were pooled, and 150 bp paired-end sequencing was performed on the Novaseq X Plus sequencer.
Banerjee R., Ajithkumar P., Keestra N., Smith J., Gimenez G., Rodger E.J., Eccles M.R., Antony J., Weeks R.J, & Chatterjee A. (2024). Targeted DNA Methylation Editing Using an All-in-One System Establishes Paradoxical Activation of EBF3. Cancers, 16(5), 898.
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8

Metagenomic Profiling of Gut Microbiome

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Shotgun DNA metagenomics sequencing was carried out on fecal DNA samples at Novogene Corp Inc. (Beijing, China). DNA samples were assessed using Qubit Fluorimeter while agarose gel electrophoresis was carried out to check for DNA degradation. The NEBNext UltraTMII DNA Library PrepKit for Illumina® (Ipswich, MA) was used to prepare the sequencing libraries according to the manufacturer’s protocols and Illumina 150 bp paired end (PE150) sequencing was conducted on the HiSeqX instruments with a targeted depth of 20 million. Quality filtering (e.g., removal of adapters, human sequences, low-quality sequences) was conducted using KneadData version 5.1 (http://huttenhower.sph.harvard.edu/kneaddata) as previously described.33 (link) All de-multiplexed, paired-end shotgun DNA metagenomic sequencing data along with the metadata were deposited into the Sequence Read Archive under BioProject accession number PRJNA739170. Reads corresponding to BA metabolizing enzymes were identified by a translated search with DIAMOND 40 (link) and a manually curated protein sequence database, respectively at two threshold levels: 1.) 80% identity over 80% sequence and 2.) 50% identity over 80% sequence. Reads corresponding to antibiotic resistance genes were identified using the Comprehensive Antimicrobial Resistance Database (CARD).
Ramos R.J., Zhu C., Joseph D.F., Thaker S., Lacomb J.F., Markarian K., Lee H.J., Petrov J.C., Monzur F., Buscaglia J.M., Chawla A., Small-Harary L., Gathungu G., Morganstern J.A., Yang J., Li J., Pamer E.G., Robertson C.E., Frank D.N., Cross J.R, & Li E. (2022). Metagenomic and bile acid metabolomic analysis of fecal microbiota transplantation for recurrent Clostridiodes difficile and/or inflammatory bowel diseases. Medical research archives, 10(10), 10.18103/mra.v10i10.3318.
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9

CTCF Binding Site Profiling by ChIC/CUT&RUN

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ChIC/CUT&RUN kit (EpiCypher Cat# 15–1016) was used to profile the CTCF binding sites, with 500,000 cells.19 (link) NEBNext® UltraTM II DNA Library Prep Kit for Illumina (Ilumina Cat# E7645S) was used to make the sequencing libraries. Sequencing was performed using Illumina NovaSeq platform for 150bp paired-end reads. Sequencing data was evaluated by FastQC and the adaptor sequence was trimmed using Trimmomatic (v0.39), followed by Bowtie2 alignment and removal of PCR duplicates by GATK (v4.2.3.0), MACS2 software was used for the narrow peak calling with the matched IgG control with the parameter ‘-p 0.05’.20 (link)
Ka-Yue Chow L., Lai-Shun Chung D., Tao L., Chan K.F., Tung S.Y., Cheong Ngan R.K., Ng W.T., Wing-Mui Lee A., Yau C.C., Lai-Wan Kwong D., Ho-Fun Lee V., Lam K.O., Liu J., Chen H., Dai W, & Lung M.L. (2022). Epigenomic landscape study reveals molecular subtypes and EBV-associated regulatory epigenome reprogramming in nasopharyngeal carcinoma. eBioMedicine, 86, 104357.
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10

Bacterial Phage Resistance Profiling

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Three independent cultures of V. alginolyticus in logarithmic phase were used to coculture with phage BUCT549, and after observing the reclouding of the culture, the surviving bacteria were used to delineate and pick single colonies for liquid culture, and tolerance to BUCT549 was confirmed after a spot assay and DAL assay. Bacteria nucleic acids were extracted using the Bacteria Genomic DNA Kit (cwbiotech) and the DNA library was prepared with the NEBNext® UltraTM II DNA Library Prep Kit for Illumina high-throughput genome sequencing.
Li J., Tian F., Hu Y., Lin W., Liu Y., Zhao F., Ren H., Pan Q., Shi T, & Tong Y. (2021). Characterization and Genomic Analysis of BUCT549, a Novel Bacteriophage Infecting Vibrio alginolyticus With Flagella as Receptor. Frontiers in Microbiology, 12, 668319.
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