The genomic DNAs were isolated from the cells using the PuregeneTM DNA purification kit (Qiagen). Library construction and exome enrichment were performed using Illumina TruSeq DNA Sample Prep Kit-Set A, SeqCap EZ Human Exome Library v2.0, and SeqCap EZ hyb and wash kit (Roche NimbleGen). Next, exome enriched libraries were sequenced using Illumina Genome Analyzer IIx. The resulting sequencing reads were aligned on human reference genome 19 (hg19) with Burrows Wheelers Aligner (BWA)34 (link). VarScan 2 was used to identify genetic variants by comparing genome sequences with hg1935 (link) and dbNSFP was used to annotate protein coding variants36 (link). Identified genetic variants with more than 10X coverage were selected for subsequent analyses. Total RNAs were extracted using RNeasy® Mini kit (Qiagen). Libraries were constructed using Illumina TruSeq RNA Sample Prep Kit v2-Set A and sequencing was carried out using the Illumina Genome Analyzer IIx. Following this, the reads were mapped on hg19 using TopHat v2.0.637 (link), and the expression level of transcripts was estimated using our customized Python code, which calculates the reads per kilobase per million reads (RPKM).
Genome analyzer iix
Manufactured by Illumina
Sourced in United States, Germany, China
The Genome Analyzer IIx is a high-throughput DNA sequencing system developed by Illumina. It is designed to perform massively parallel sequencing of DNA samples, allowing for rapid and accurate determination of genetic sequences.
Automatically generated - may contain errors
Lab products found in correlation
Hiseq 2000, by Illumina (66 mentions)
Agilent 2100 bioanalyzer, by Agilent Technologies (40 mentions)
Rneasy mini kit, by Qiagen (21 mentions)
Hiseq 2500, by Illumina (20 mentions)
Trizol reagent, by Thermo Fisher Scientific (19 mentions)
2100 bioanalyzer, by Agilent Technologies (16 mentions)
Truseq rna sample prep kit, by Illumina (12 mentions)
Truseq rna sample preparation kit, by Illumina (12 mentions)
Trizol, by Thermo Fisher Scientific (11 mentions)
Superscript 2 reverse transcriptase, by Thermo Fisher Scientific (8 mentions)
Mrna seq sample prep kit, by Illumina (8 mentions)
Truseq rna sample prep kit v2, by Illumina (8 mentions)
Truseq small rna sample prep kit, by Illumina (7 mentions)
Mrna seq sample preparation kit, by Illumina (7 mentions)
Truseq rna sample preparation kit v2, by Illumina (7 mentions)
Cluster station, by Illumina (6 mentions)
Nextseq 500, by Illumina (6 mentions)
Truseq sbs kit v5, by Illumina (6 mentions)
Flow cell, by Illumina (6 mentions)
Nebnext poly a mrna magnetic isolation module, by New England Biolabs (5 mentions)
497 protocols using genome analyzer iix
1
Whole-Exome and Transcriptome Sequencing
2
Transcriptome and Small RNA Profiling of P. kurrooa
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
P. kurrooa plants were kept at 15 °C and 25 °C in two separate plant growth chambers (350 μmol m−2 s−1 photosynthetic photon flux density; 16 h photoperiod) for 2 weeks followed by sampling of leaf and rhizome tissues at both the temperatures. RNA was isolated as described by Ghawana et al.90 (link) and its integrity and concentration was checked by using RNA Nano chip on Agilent Bioanalyzer 2100 (Agilent technologies, USA). cDNA and small RNA libraries were prepared using truseq RNA library preparation kit (Illumina, USA) and truseq small RNA sample preparation kit (Illumina, USA), respectively, following the manufacturer’s instructions. Paired end (PE) 36 × 2 bp sequencing of cDNA libraries was carried out on Illumina Genome Analyzer IIx (Illumina, USA) as per the manufacturer’s instructions. Sequencing of small RNA libraries was also carried out on Illumina Genome Analyzer IIx (Illumina, USA) as per the manufacturer’s instructions.
3
Mutmap Methodology for Gene Mapping
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Mutmap [57 (link)] methodology was used for gene mapping. Briefly, four F1 and four F2 populations derived from the crosses WT×sg2-1, sg2-1×WT, WT×sg2-2, and sg2-2×WT were used for genetic analysis. F2 populations derived from the crosses 02428 × sg2-1 and 02428 × sg2-2 were used for mapping of the mutant gene. DNA from 25 BC1F2 plants with small grain phenotype similar to sg2-1 was extracted and pooled in equal proportions, and subjected to whole-genome sequencing using Illumina Genome Analyzer IIx (Novogene, Beijing, China). Mixed DNA (5 μg) was used for preparation of libraries for Illumina sequencing according to the protocol for the Paired-End DNA Sample Prep kit (Novogene, Beijing, China). The libraries were used for cluster generation on a flow cell and sequenced for 76 cycles on an Illumina Genome Analyzer IIx. DNA from WT was re-sequenced as a control. The SNPs/INDELs indexes were calculated as previously described [57 (link)]. Sequences of the PCR primers used for mapping and the amplified sg2-2 mutant genomic DNA sequence are given in Table S5 .
4
Gene Expression Profiling of ESCC
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The publically accessible GEO databank, a genomics data repository for next-generation sequencing and comprising sequence- and array-based data of gene profiles, was used for profiling studies of ESCC gene expression. Three datasets GSE130078, GSE32424, GSE29968 and were picked out of the GEO repository. Three tissues of ESCC and three typical tissues of the esophagus were a part of the GSE29968 dataset, which was obtained from the GPL10999 Illumina Genome Analyzer IIx. The GSE32424 dataset includes 7 ESCC tissues and 5 normal tissues of the esophagus and was derived from the GPL10999 Illumina Genome Analyzer IIx. The dataset GSE130078 includes 23 ESCC tissues and 23 normal tissues of the esophagus and was generated from the GPL11154 Illumina HiSeq 2000. The various steps of the research were conducted in line with the Helsinki Declaration (2013 revision).
5
Transcriptome Assembly and Sequencing for Marine Species
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Because there are no reference transcriptomes for Pacific abalone and sand worms, we conducted paired-end RNA-seq on the two species and assembled their transcriptomes. The transcriptome of the Pacific abalone was sequenced using the Illumina Genome Analyzer IIx platform at Biomarker Technologies CO., LTD, while that of the sand worm was sequenced using the Illumina HiSeq 2000 platform at BGI-Shenzhen. RNAs from each developmental stage were first mixed in equal quantities. Then, sequencing libraries were constructed according to the Illumina’s recommended protocol.
Single-end RNA-seq was conducted to obtain the number of gene reads for each developmental stage of the three species. Single-end RNA sequencing data for the oyster and abalone samples was generated using the Illumina Cluster Station and Illumina Genome Analyzer IIx, while the sand worm was sequenced using the Illumina HiSeq 2000 platform. The sequencing libraries were constructed according to Illumina’s recommended protocol.
A short insert library with 300-bp fragments was constructed for resequencing of the oyster genome using a standard protocol recommended by Illumina. The DNA was sequenced using the Illumina HiSeq 2000 platform.
Single-end RNA-seq was conducted to obtain the number of gene reads for each developmental stage of the three species. Single-end RNA sequencing data for the oyster and abalone samples was generated using the Illumina Cluster Station and Illumina Genome Analyzer IIx, while the sand worm was sequenced using the Illumina HiSeq 2000 platform. The sequencing libraries were constructed according to Illumina’s recommended protocol.
A short insert library with 300-bp fragments was constructed for resequencing of the oyster genome using a standard protocol recommended by Illumina. The DNA was sequenced using the Illumina HiSeq 2000 platform.
6
Drosophila Transcriptome Sequencing Methods
7
Next-Generation Sequencing of Diverse Libraries
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The 33 libraries were sequenced on 3 different lanes. The first run including 19 libraries was processed on an Illumina Genome Analyzer IIx using 72 bp single-end reads. The second run of four libraries was also processed on an Illumina Genome Analyzer IIx but using 72 bp paired-end reads. These first two runs were subcontracted to Ambry Genetics (Aliso Viejo, CA, USA). The third run of 10 libraries was run on an Illumina HiSeq 2500 instrument with 100 bp paired-end reads at the Donnelly Sequencing Centre of the University of Toronto (Canada). Initial data processing and base calling, including extraction of cluster intensities, was done using RTA1.8 (SCS version 2.8). Sequence quality filtering script was executed in the Illumina CASAVA software (ver 1.7.0, Illumina, Hayward, CA).
8
Drosophila Transcriptome Sequencing Methods
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Libraries were generated and sequenced on an Illumina Genome Analyzer IIx or HiSeq 2000 using paired-end chemistry and 76 or 100bp cycles. 454 sequencing used poly(A)+ RNA from Oregon R adult males and females and mixed-staged y1 cn1 bw1 sp1 embryos. Sequences are available from the Short Read Archive and the modENCODE website (http://www.modencode.org /). CAGE46 (link) was sequenced on a Illumina Genome Analyzer IIx with 36bp reads. Poly(A)+seq was generated using a custom protocol (Supplementary Methods ).
9
RNA-Seq Analysis of Blood Transcriptome
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The mRNA obtained from blood was sequenced using a Genome Analyzer IIx from Illumina. Total RNA was extracted from peripheral blood of each individual. The mRNA was isolated from the total RNA and, once transformed into cDNA, was fragmented by sonication. Fragments of 300bp on average were selected to construct the libraries for sequencing. The libraries to sequence were constructed using the reagents according to the indications of the manufacturer (Illumina). Single-end sequences of 35 nucleotides were produced using a Genome Analyzer IIx (Illumina). Alignment of the reads was performed in an Simple Linux Utility for Resource Management High Performance Computer server running Tophat 2.0.6 with default options (Trapnell et al., 2009 (link)). Tophat aligns RNA-Seq reads to genomes using the Bowtie 2.0.2 alignment program (Langmead et al., 2009 (link)), and then analyzes the mapping results to identify splice junctions between exons.
10
ESCC Gene Expression Profiling Using GEO Datasets
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
ESCC gene expression profiling studies were carried out using the GEO database. Three GEO datasets GSE29968, GSE32424 and GSE130078 were downloaded. GSE29968 dataset was based on the GPL10999 Illumina Genome Analyzer IIx and included 3 normal esophageal tissues and 3 ESCC tissues. GSE32424 dataset was based on the GPL10999 Illumina Genome Analyzer IIx and included 5 normal esophageal tissues and 7 ESCC tissues. GSE130078 dataset was based on the GPL11154 Illumina HiSeq 2000 and included 23 normal esophageal tissues and 23 ESCC tissues.
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!