Genome libraries were independently prepared for the parental lines and the four F20 pigmentation subgroups for each cross. For each library, genomic DNA was extracted from a pool of 30 females by chloroform extraction and ethanol precipitation. DNA was then fragmented using the Bioruptor sonicator (Diagenode), and paired-end libraries with approximately 300-bp inserts were prepared using the NEBNext DNA Library Prep Reagent Set for Illumina (New England Biolabs no. E6000L). Library concentration and quality were assessed using an Agilent 2100 Bioanalyzer (Agilent Technologies) and were sequenced at the UW–Madison Biotechnology Center on the Illumina HiSequation 2000 platform with 100-bp paired read lengths.
Hisequation 2000 platform
Manufactured by Illumina
The HiSequation 2000 platform is a high-throughput DNA sequencing system designed for a wide range of applications. It utilizes sequencing-by-synthesis technology to generate high-quality sequence data. The platform offers scalable throughput and flexible configuration options to suit various research and clinical needs.
Automatically generated - may contain errors
Lab products found in correlation
Agilent 2100 bioanalyzer, by Agilent Technologies (1 mentions)
Bioruptor sonicator, by Diagenode (1 mentions)
Trizol rna extraction kit, by Thermo Fisher Scientific (1 mentions)
Nebnext ultra rna library prep kit, by New England Biolabs (1 mentions)
Truseq pe cluster kit v3 cbot hs, by Illumina (1 mentions)
Rna nano 6000 assay kit, by Agilent Technologies (1 mentions)
Agilent bioanalyzer 2100 system, by Agilent Technologies (1 mentions)
Truseq rna sample prep kit version 2, by Illumina (1 mentions)
Trizol buffer, by Takara Bio (1 mentions)
4 protocols using hisequation 2000 platform
1
Genome Sequencing of Pigmentation Subgroups
2
High-throughput RNA Sequencing of Larval Heads
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Total RNA was extracted from pools of five larval heads using a TRIzol RNA extraction kit (Invitrogen, Carlsbad, CA). RNA concentration and integrity were determined by an RNA Nano 6000 Assay Kit and the Agilent Bioanalyzer 2100 system (Agilent Technologies, Santa Clara, CA). The isolated RNA pellets were stored at −80° until needed. Two biological replicates were used in the analysis. Six RNA-Seq libraries were generated using a NEBNextUltra RNA Library Prep Kit (NEB) following the manufacturer’s recommendations. Clustering of the index-coded samples was performed on a cBot Cluster Generation System using a TruSeq PE Cluster Kit v3-cBot-HS (Illumina) according to the manufacturer’s instructions. After cluster generation, the library preparations were sequenced on an Illumina HiSequation 2000 platform, and paired-end reads were generated (read length = 150 bp) (Illumina Inc., San Diego, CA).
3
RNA-seq Transcriptome Profiling of OGD Groups
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RNA-seq transcriptome array of the non-OGD, OGD, and OGD + PDRN groups was performed at Macrogen Inc. (Seoul, Korea) with the HiSequation 2000 platform (Illumina, San Diego, CA, USA) according to methods detailed in our previous study20 (link).
4
Dose-dependent RNA-seq of OVA-stimulated DCs
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On the sixth day, OVA solutions of different doses were added to the DCs taken from mouse bone marrow, and the concentration of OVA in each hole was 0, 10, 100, and 10,000μg /ml, respectively. Three samples were tested for each group, and the culture lasted for 24 hours. After the collection of suspension cells, the cells, in each group, were separated and resuscitated in 0.5ml Trizol buffer (Takara, Dalian, China). RNA-seq was performed (Wuhan Kangce Co., LTD., China). Trizol extraction is a reliable method for obtaining high-quality total RNA. RNA-seq library was constructed by Illumina TruSeq RNA Sample Prep Kit version 2 (Illumina, San Diego, CA) and RNA was sequenced by Illumina HiSEquation 2000 platform (Illumina). Intergroup analysis: 10-0 DCs (10 DCs group compare with 0 DCs group), 100-0 DCs (100 DCs group compare with 0 DCs group), 10000-0 DCs (1000 DCs group compare with 0 DCs group).
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