150 cycle mid output kit

Manufactured by Illumina

Sourced in United States

The 150-cycle mid output kit is a lab equipment product that provides a sequencing solution for Illumina's sequencing platforms. It is designed to enable up to 150 sequencing cycles, allowing for increased read lengths and higher data output. The kit includes all the necessary reagents and consumables required for the sequencing process.

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Lab products found in correlation

Nextera xt index kit, by Illumina (2 mentions) Qubit fluorometer, by Thermo Fisher Scientific (2 mentions) Nextseq 500 platform, by Illumina (1 mentions) Kapa library quantification kit, by Roche (1 mentions) Q5 high fidelity dna polymerase, by New England Biolabs (1 mentions) Ampure xp bead, by Beckman Coulter (1 mentions) Kapa hifi hotstart readymix, by Illumina (1 mentions) 4200 tapestation system, by Agilent Technologies (1 mentions) Qubit dsdna high sensitivity assay, by Thermo Fisher Scientific (1 mentions) D1000 screentape kit, by Agilent Technologies (1 mentions)

Spelling variants of this product

Mid Output Kit (15 citations) NextSeq 500/550 Mid Output Kit v2.5 (150 Cycles) (3 citations)

3 protocols using 150 cycle mid output kit

Optimized CROP-seq gRNA Library Preparation

Cited 1 time

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CROP-seq-opti gRNA sequencing libraries were prepared as previously described [113 (link)]. Briefly, 100ng of each gDNA sample was PCR amplified in triplicate with Q5 High-Fidelity DNA Polymerase (NEB #M0494S) with 500nM primers (S4 Table) flanking the guide sequence cassette and including Illumina adaptor sequences and sample index sequences (98°C x 30s, 98°C x 10s, 72°C x 45s, 25 cycles). PCR products were purified using 2.0X AMPure XP magnetic beads (Beckman Coulter #A63881) according to manufacturer’s protocol. Sequencing libraries were quantified with the KAPA Library Quantification Kit (Roche #07960140001), pooled, and sequenced in multiplex on the Illumina NextSeq 500 platform using a 150-cycle mid output kit (Illumina # 20024904) with read configuration of 167 bases (read 1) and 8 bases (i7 index).

Danziger O., Patel R.S., DeGrace E.J., Rosen M.R, & Rosenberg B.R. (2022). Inducible CRISPR activation screen for interferon-stimulated genes identifies OAS1 as a SARS-CoV-2 restriction factor. PLoS Pathogens, 18(4), e1010464.

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Amplicon Sequencing Library Preparation

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The multiplex PCR products (25 μl) were cleaned by AMPure XP beads, Beckman Coulter (X1), and eluted in 25 μl elution buffer. The purified products (7.5 μl) were subjected to a second PCR to include unique index sequences (N7XX and S5XX) for barcoding of each sample using Nextera XT Index Kit (Illumina, San Diego, CA, USA). Then, 5 μl from each barcoded sample was pooled together, mixed and spun down. Finally, 100 μl of pooled DNA was purified using X1 AMPure XP beads and quantified by Qubit® Fluorometer (Invitrogen) machine. The concentration of 4.0 nM was prepared from the pooled sample. At least 10,000 reads for each sample were targeted. Samples were deep-sequenced with the Nextseq500 machine using the 150-cycle mid output kit (Illumina, Inc., USA) from the forward read direction.

Nasereddin A., Ereqat S., Al-Jawabreh A., Taradeh M., Abbasi I., Al-Jawabreh H., Sawalha S, & Abdeen Z. (2022). Concurrent molecular characterization of sand flies and Leishmania parasites by amplicon-based next-generation sequencing. Parasites & Vectors, 15, 262.

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Indexing and Sequencing of PCR Products

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The purified PCR products of EJ5 (5 μl) were subjected to a second round of amplification to assign unique index sequences (barcode) for each sample using Nextera XT Index Kit (Illumina, San Diego, CA, USA), 2x KAPA HiFi HotStart Ready Mix (25 μl), PCR Grade water (10 μl) in total reaction volume of 50μl. The following PCR program was used: 95°C for 3 minutes, followed with 8 cycles of 95°C for 30 seconds, 55°C for 30 seconds and 72°C for 30 seconds, then 72°C for 5 minutes and finally hold at 4°C. The product was purified by Agencourt AMPure XP system (X1) (A63881; Beckman Coulter Genomics), and eluted in 30 μl elution buffer. Library purity and quantity were evaluated by 4200 TapeStation System (Agilent Technologies, Inc., Santa Clara, CA, USA) using D1000 ScreenTape kit (Agilent) and by Qubit® Fluorometer (Invitrogen, Carlsbad, CA, USA) using Qubit dsDNA high-sensitivity assay (Invitrogen). Pool concentration of 4 nM was prepared and 1.5×10⁶ or 3×10⁵ reads for each sample were targeted. Samples were deep sequenced on NextSeq 500/550 machine using the 150-cycle Mid Output Kit (Illumina).

Britan-Rosich Y., Ma J., Kotler E., Hassan F., Botvinnik A., Smith Y., Moshel O., Nasereddin A., Sharma G., Pikarsky E., Ross S, & Kotler M. (2022). APOBEC3G protects the genome of human cultured cells and mice from radiation-induced damage. The FEBS journal, 290(7), 1822-1839.

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