High sensitivity bioanalyzer kit

Manufactured by Agilent Technologies

Sourced in United States

The High sensitivity Bioanalyzer kit is a lab equipment product from Agilent Technologies designed for high-sensitivity analysis of nucleic acid samples. The kit provides an automated, microfluidics-based platform for sizing and quantification of DNA, RNA, and protein samples.

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Lab products found in correlation

Quant it picogreen dsdna kit, by Thermo Fisher Scientific (1 mentions) Dna mini kit, by Qiagen (1 mentions) Truseq nano dna kit, by Illumina (1 mentions) Bioruptor, by Diagenode (1 mentions) Nextseq 500, by Illumina (1 mentions) Eb buffer, by Qiagen (1 mentions) Ampure xp magnetic beads, by Qiagen (1 mentions) Bioanalyzer high sensitivity chip, by Agilent Technologies (1 mentions) Qubit dsdna hs kit, by Thermo Fisher Scientific (1 mentions) Miseq sequencing, by Illumina (1 mentions) Nebnext multiplex small rna library prep kit, by New England Biolabs (1 mentions) Nebnext magnesium rna fragmentation module kit, by New England Biolabs (1 mentions) Nextseq 500 platform, by Illumina (1 mentions)

5 protocols using high sensitivity bioanalyzer kit

Mapping Transgene Insertion Sites

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We performed whole-genome sequencing to identify the insertion-sites of transgene in the mouse genome. Paired-end 150 bp-long sequencing reads were aligned to a modified version of the reference genome (GRCm38/mm10) using the bowtie aligner (bowtie-bio.sourceforge.net). The reference was modified to include a copy of the original cloning vector. Sequencing reads that uniquely mapped both to the mouse genome and to the cloning vector sequence were used to localize the insertion sites. Raw-sequencing reads in the form of.fastq files were deposited in the European Nucleotide Archive with the accession (E-MTAB-5502). To generate the library, DNA was extracted from liver using the DNA Mini kit (Qiagen) and fragmented using Bioruptor (Diagenode). Briefly, 380 ng DNA in 100 microL in 500 microL tubes were subjected to sonication with the following parameters: power setting low, cycle 30 sec on/90 sec off, time 8 minutes in ice-cold, degassed water, resulting in average 550 bp size fragments. The library was prepared using the TruSeq Nano DNA kit (Illumina). DNA and library quantifications were performed with the Quant-iT PicoGreen dsDNA Kit (Invitrogen). Library quality was assessed by using the High sensitivity Bioanalyzer kit (Agilent).

Barzago M.M., Kurosaki M., Fratelli M., Bolis M., Giudice C., Nordio L., Cerri E., Domenici L., Terao M, & Garattini E. (2017). Generation of a new mouse model of glaucoma characterized by reduced expression of the AP-2β and AP-2δ proteins. Scientific Reports, 7, 11140.

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Sequencing Enriched DNA Libraries

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Libraries were quantified between the range of 200bp and 1 kbp on a High Sensitivity Bioanalyzer kit (Agilent, Cat. 5067-4626). Libraries were sequenced on an Illumina NextSeq^® 500 loaded at 0.8 pM with a custom sequencing chemistry protocol (Read 1: 50 imaged cycles; Index Read 1: 8 imaged cycles, 27 dark cycles, 10 imaged cycles; Index Read 2: 8 imaged cycles, 21 dark cycles, 10 imaged cycles; Read 2: 50 imaged cycles) using custom sequencing primers described in Amini et. al. 2015, Ref.14 (link). QRP and DOP libraries were sequenced using standard primers on the NextSeq^® 500 using high-capacity 75 cycle kits with dual-indexing. For QRP there is an additional challenge that the first 15 bp of the read are highly enriched for “G” bases, which are non-fluorescent with the NextSeq^® 2-color chemistry and therefore cluster identification on the instrument fails. We therefore sequenced the libraries using a custom sequencing protocol that skips this region (Read 1: 15 dark cycles, 50 imaged cycles; Index Read 1: 10 imaged cycles; Index Read 2: 10 imaged cycles).

Vitak S.A., Torkenczy K.A., Rosenkrantz J.L., Fields A.J., Christiansen L., Wong M.H., Carbone L., Steemers F.J, & Adey A. (2017). Sequencing thousands of single-cell genomes with combinatorial indexing. Nature methods, 14(3), 302-308.

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Single-cell RNA-seq cDNA amplification protocol

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5.7 μl of retro‐transcription mix (see reference for details) was added to each sample. Retro‐transcription was carried out according to the original Smart‐seq2 protocol and cDNA was then pre‐amplified for 15 cycles (Picelli et al, 2014 (link)). After PCR pre‐amplification, cDNA was purified using Ampure XP magnetic beads according to the manufacturer's instructions, in a ratio of 0.8 to 1 with cDNA, resuspended in 17.5 μl of buffer EB (Qiagen) and stored at −20°C. Quality and concentration of the cDNA generated were assessed using High‐Sensitivity Bioanalyzer kit (Agilent).

Kaeppler J.R., Chen J., Buono M., Vermeer J., Kannan P., Cheng W., Voukantsis D., Thompson J.M., Hill M.A., Allen D., Gomes A., Kersemans V., Kinchesh P., Smart S., Buffa F., Nerlov C., Muschel R.J, & Markelc B. (2022). Endothelial cell death after ionizing radiation does not impair vascular structure in mouse tumor models. EMBO Reports, 23(9), e53221.

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Illumina MiSeq Sequencing Library Preparation

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Library preparation was performed at the Genotypic Technology's Genomics facility. The PCR amplified product was checked on an agarose gel before proceeding to PCR indexing. The PCR index reaction was conducted using a Nextera® XT Index Kit v2 Set D kit from which Illumina sequencing adapters and dual indexing barcodes were added using limited cycle PCR (Initial Denaturation 95 °C for 5min, Followed by 10 cycles of 98 °C for 20s, 64 °C for 15s, 72 °C for 35s, a final extension of 72 °C for 5min). This provided a final product of ∼520bp and ∼570bp. The library was cleaned using HighPrep PCR magnetic beads (HighPrep PCR, Magbio, Switzerland) and was Qubit quantified using Qubit ds DNA HS kit (Invitrogen, USA). Qubit quantification resulted in concentration of 1.91 ng/μl with total yield of 19.1 ng. The prepared library was then validated for quality using High Sensitivity Bioanalyzer Kit (Agilent Technologies, USA) by running an aliquot on High Sensitivity Bioanalyzer Chip (Agilent Technologies, USA). The library was then pooled for Illumina MiSeq sequencing using the 250bp paired end read chemistry.

Ragupathy S., Faller A.C., Shanmughanandhan D., Kesanakurti P., Shaanker R.U., Ravikanth G., Sathishkumar R., Mathivanan N., Song J., Han J, & Newmaster S. (2019). Exploring DNA quantity and quality from raw materials to botanical extracts. Heliyon, 5(6), e01935.

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RNA-seq Library Preparation for Microbiome

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A 5-mL sample of the culture was taken from at least two chemostat experiments with 4 replicates in total and mixed with RNAlater, which was incubated overnight at 4°C and then stored at −80°C. RNA-seq libraries were prepared as previously described for in vitro samples (5 (link)). rRNA was depleted using the MICROBExpress bacterial mRNA enrichment kit (Sigma). The depleted RNA was fragmented for 2 min with the NEBNext magnesium RNA fragmentation module kit, and cDNA libraries were prepared using the NEBNext multiplex small RNA library prep kit (New England BioLabs). The final RNA concentration for each sample was assessed with a Qubit dsDNA HS (double-stranded DNA high-sensitivity) assay kit (Thermo) and the high-sensitivity Bioanalyzer kit (Agilent). Libraries were sequenced at the Molecular Evolution Core at the Georgia Institute of Technology on an Illumina NextSeq500 platform using 75-bp single-end runs.

Zhang M., Michie K.L., Cornforth D.M., Dolan S.K., Wang Y, & Whiteley M. (2022). Impact of Growth Rate on the Protein-mRNA Ratio in Pseudomonas aeruginosa. mBio, 14(1), e03067-22.

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