Structured Review

Illumina Inc hiseq 2500
Zinc-related and gli cluster genes were upregulated during phagocytosis. ( A ) Total RNA was extracted from the macrophage phagocytic cells infected with conidia of A. fumigatus , and libraries were prepared from 2 µg of total RNA. High-throughput sequencing was performed as paired-end 100 bp sequencing using a <t>HiSeq</t> 2500 (Illumina, Inc., USA). mRNA-seq reads were mapped using the TopHat software tool to obtain an alignment file. ( B ) Total RNA was extracted from the phagocytic cells infected with conidia after 0 and 1 h postinfection, and cDNA was synthesized using a TOPscript™ cDNA synthesis kit (Enzynomics). Then, qRT-PCR was performed using KAPA SYBR ® for LightCycler ® 480 with the primer sets for the indicated genes. ( C ) The transcriptomic profile during phagocytosis is shown. The gene expression profile was grouped into zinc, copper, iron, and gliotoxin metabolism. Upregulated genes were found in all categories, and ZafA and GliZ were upregulated during phagocytosis. Up- and downregulated genes are colored to show fold change. * p
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Images

1) Product Images from "The Role of Zinc in Gliotoxin Biosynthesis of Aspergillus fumigatus"

Article Title: The Role of Zinc in Gliotoxin Biosynthesis of Aspergillus fumigatus

Journal: International Journal of Molecular Sciences

doi: 10.3390/ijms20246192

Zinc-related and gli cluster genes were upregulated during phagocytosis. ( A ) Total RNA was extracted from the macrophage phagocytic cells infected with conidia of A. fumigatus , and libraries were prepared from 2 µg of total RNA. High-throughput sequencing was performed as paired-end 100 bp sequencing using a HiSeq 2500 (Illumina, Inc., USA). mRNA-seq reads were mapped using the TopHat software tool to obtain an alignment file. ( B ) Total RNA was extracted from the phagocytic cells infected with conidia after 0 and 1 h postinfection, and cDNA was synthesized using a TOPscript™ cDNA synthesis kit (Enzynomics). Then, qRT-PCR was performed using KAPA SYBR ® for LightCycler ® 480 with the primer sets for the indicated genes. ( C ) The transcriptomic profile during phagocytosis is shown. The gene expression profile was grouped into zinc, copper, iron, and gliotoxin metabolism. Upregulated genes were found in all categories, and ZafA and GliZ were upregulated during phagocytosis. Up- and downregulated genes are colored to show fold change. * p
Figure Legend Snippet: Zinc-related and gli cluster genes were upregulated during phagocytosis. ( A ) Total RNA was extracted from the macrophage phagocytic cells infected with conidia of A. fumigatus , and libraries were prepared from 2 µg of total RNA. High-throughput sequencing was performed as paired-end 100 bp sequencing using a HiSeq 2500 (Illumina, Inc., USA). mRNA-seq reads were mapped using the TopHat software tool to obtain an alignment file. ( B ) Total RNA was extracted from the phagocytic cells infected with conidia after 0 and 1 h postinfection, and cDNA was synthesized using a TOPscript™ cDNA synthesis kit (Enzynomics). Then, qRT-PCR was performed using KAPA SYBR ® for LightCycler ® 480 with the primer sets for the indicated genes. ( C ) The transcriptomic profile during phagocytosis is shown. The gene expression profile was grouped into zinc, copper, iron, and gliotoxin metabolism. Upregulated genes were found in all categories, and ZafA and GliZ were upregulated during phagocytosis. Up- and downregulated genes are colored to show fold change. * p

Techniques Used: Infection, Next-Generation Sequencing, Sequencing, Software, Synthesized, Quantitative RT-PCR, Expressing

2) Product Images from "Integrating reductive and synthetic approaches in biology using man-made cell-like compartments"

Article Title: Integrating reductive and synthetic approaches in biology using man-made cell-like compartments

Journal: Scientific Reports

doi: 10.1038/srep04722

(a) Hierarchical cluster analysis for genes included in non-fluorescent liposomes. Ten non-fluorescent liposomes that did not show β-galactoside hydrolysis activity were isolated, and the genes included in these liposomes were determined by multiplex next-generation sequencing using HiSeq 2500. In the hierarchical cluster analysis for these genes, there was no commonality among the genes included in each liposome, which indicated a random pattern. Each row represents a separate gene, each column represents a separate liposome and genes found in each liposome are shown in red. (b) Hierarchical cluster analysis for the genes included in fluorescent liposomes. Hierarchical cluster analysis revealed that  LacZ  was a clear cluster that was included in all fluorescent liposomes and with no other common factors.
Figure Legend Snippet: (a) Hierarchical cluster analysis for genes included in non-fluorescent liposomes. Ten non-fluorescent liposomes that did not show β-galactoside hydrolysis activity were isolated, and the genes included in these liposomes were determined by multiplex next-generation sequencing using HiSeq 2500. In the hierarchical cluster analysis for these genes, there was no commonality among the genes included in each liposome, which indicated a random pattern. Each row represents a separate gene, each column represents a separate liposome and genes found in each liposome are shown in red. (b) Hierarchical cluster analysis for the genes included in fluorescent liposomes. Hierarchical cluster analysis revealed that LacZ was a clear cluster that was included in all fluorescent liposomes and with no other common factors.

Techniques Used: Activity Assay, Isolation, Multiplex Assay, Next-Generation Sequencing

3) Product Images from "Larval application of sodium channel homologous dsRNA restores pyrethroid insecticide susceptibility in a resistant adult mosquito population"

Article Title: Larval application of sodium channel homologous dsRNA restores pyrethroid insecticide susceptibility in a resistant adult mosquito population

Journal: Parasites & Vectors

doi: 10.1186/s13071-016-1634-y

VGSC small RNAs persist until the adult stage of  Ae. aegypti  treated by dsRNA-VGSC during larval stage.  a  Small RNA libraries were constructed from 5 day-old adult mosquitoes previously soaked in VGSC 422  dsRNA or Random dsRNA (0.5 μg/μl) and sequenced using Hiseq 2500. Data are presented as the number of reads normalized to one million mapped reads per sample (RPM). Forward strand (sense) reads are positive RPM values (depicted in  blue ); reverse strand (anti-sense) reads are in negative RPM values (depicted in  red ). Positions for which RPM values are  >  = 10 RPM were considered expressed.  b  Size distribution of small RNAs matching the VGSC422 transcript (Accession no. KC107440.1). nt: nucleotides
Figure Legend Snippet: VGSC small RNAs persist until the adult stage of Ae. aegypti treated by dsRNA-VGSC during larval stage. a Small RNA libraries were constructed from 5 day-old adult mosquitoes previously soaked in VGSC 422 dsRNA or Random dsRNA (0.5 μg/μl) and sequenced using Hiseq 2500. Data are presented as the number of reads normalized to one million mapped reads per sample (RPM). Forward strand (sense) reads are positive RPM values (depicted in blue ); reverse strand (anti-sense) reads are in negative RPM values (depicted in red ). Positions for which RPM values are  >  = 10 RPM were considered expressed. b Size distribution of small RNAs matching the VGSC422 transcript (Accession no. KC107440.1). nt: nucleotides

Techniques Used: Construct

4) Product Images from "Integrating reductive and synthetic approaches in biology using man-made cell-like compartments"

Article Title: Integrating reductive and synthetic approaches in biology using man-made cell-like compartments

Journal: Scientific Reports

doi: 10.1038/srep04722

(a) Hierarchical cluster analysis for genes included in non-fluorescent liposomes. Ten non-fluorescent liposomes that did not show β-galactoside hydrolysis activity were isolated, and the genes included in these liposomes were determined by multiplex next-generation sequencing using HiSeq 2500. In the hierarchical cluster analysis for these genes, there was no commonality among the genes included in each liposome, which indicated a random pattern. Each row represents a separate gene, each column represents a separate liposome and genes found in each liposome are shown in red. (b) Hierarchical cluster analysis for the genes included in fluorescent liposomes. Hierarchical cluster analysis revealed that  LacZ  was a clear cluster that was included in all fluorescent liposomes and with no other common factors.
Figure Legend Snippet: (a) Hierarchical cluster analysis for genes included in non-fluorescent liposomes. Ten non-fluorescent liposomes that did not show β-galactoside hydrolysis activity were isolated, and the genes included in these liposomes were determined by multiplex next-generation sequencing using HiSeq 2500. In the hierarchical cluster analysis for these genes, there was no commonality among the genes included in each liposome, which indicated a random pattern. Each row represents a separate gene, each column represents a separate liposome and genes found in each liposome are shown in red. (b) Hierarchical cluster analysis for the genes included in fluorescent liposomes. Hierarchical cluster analysis revealed that LacZ was a clear cluster that was included in all fluorescent liposomes and with no other common factors.

Techniques Used: Activity Assay, Isolation, Multiplex Assay, Next-Generation Sequencing

5) Product Images from "Guidelines for whole genome bisulphite sequencing of intact and FFPET DNA on the Illumina HiSeq X Ten"

Article Title: Guidelines for whole genome bisulphite sequencing of intact and FFPET DNA on the Illumina HiSeq X Ten

Journal: Epigenetics & Chromatin

doi: 10.1186/s13072-018-0194-0

Coverage comparison between HiSeq 2500 and HiSeq X Ten. a Plot shows the fraction of genome covered at different depths for four samples sequenced together on one lane of the HiSeq 2500 versus when each of the samples is sequenced on a single lane of the HiSeq X Ten. The coverage plot for the HiSeq 2500 HO mode is the merged coverage obtained from multiplexing the four samples. b Plot shows the fraction of CpG sites covered at different depths when four clinical samples are sequenced together on one lane of the HiSeq 2500 versus when each of the samples is sequenced on a single lane of the HiSeq X Ten. c , d Box plot showing the coverage distribution across exons, intergenic regions, introns, promoter regions and repeat regions of the genome for a sample sequenced on one lane of HiSeq X Ten ( c ) and HiSeq 2500 ( d )
Figure Legend Snippet: Coverage comparison between HiSeq 2500 and HiSeq X Ten. a Plot shows the fraction of genome covered at different depths for four samples sequenced together on one lane of the HiSeq 2500 versus when each of the samples is sequenced on a single lane of the HiSeq X Ten. The coverage plot for the HiSeq 2500 HO mode is the merged coverage obtained from multiplexing the four samples. b Plot shows the fraction of CpG sites covered at different depths when four clinical samples are sequenced together on one lane of the HiSeq 2500 versus when each of the samples is sequenced on a single lane of the HiSeq X Ten. c , d Box plot showing the coverage distribution across exons, intergenic regions, introns, promoter regions and repeat regions of the genome for a sample sequenced on one lane of HiSeq X Ten ( c ) and HiSeq 2500 ( d )

Techniques Used: Multiplexing

Comparison of methylation correlation between HiSeq 2500 and HiSeq X Ten. a Correlation plots of methylation levels obtained from a cell line, a clinical sample and a FFPET sample sequenced on the Hiseq 2500 versus HiSeq X Ten (Pearson r > 0.94). b Correlation of methylation values obtained from HiSeq 2500 and HiSeq X Ten for a cell line, clinical sample and a FFPET sample after grouping them into four bins of methylation percentages. c Average kappa values for six sample pairs, including two cell lines, two clinical samples and two FFPE samples compared between the HiSeq 2500 and HiSeq X Ten platform. d Bar plot showing the distribution of percentage of discordant sites across the genome for a cell line, clinical sample and a FFPET sample
Figure Legend Snippet: Comparison of methylation correlation between HiSeq 2500 and HiSeq X Ten. a Correlation plots of methylation levels obtained from a cell line, a clinical sample and a FFPET sample sequenced on the Hiseq 2500 versus HiSeq X Ten (Pearson r > 0.94). b Correlation of methylation values obtained from HiSeq 2500 and HiSeq X Ten for a cell line, clinical sample and a FFPET sample after grouping them into four bins of methylation percentages. c Average kappa values for six sample pairs, including two cell lines, two clinical samples and two FFPE samples compared between the HiSeq 2500 and HiSeq X Ten platform. d Bar plot showing the distribution of percentage of discordant sites across the genome for a cell line, clinical sample and a FFPET sample

Techniques Used: Methylation, Formalin-fixed Paraffin-Embedded

Comparing different library preparation methods using genomic DNA on the HiSeq 2500.  a  Cluster plot of sequencing output metrics obtained from different library preparation methods from intact genomic DNA.  b  Bar graph showing fragment size distribution for different bead size selection for each library preparation method.  c  Plot with  x -axis indicating coverage and  y -axis indicating fragment size illustrating how increase in fragment size leads to improved coverage
Figure Legend Snippet: Comparing different library preparation methods using genomic DNA on the HiSeq 2500. a Cluster plot of sequencing output metrics obtained from different library preparation methods from intact genomic DNA. b Bar graph showing fragment size distribution for different bead size selection for each library preparation method. c Plot with x -axis indicating coverage and y -axis indicating fragment size illustrating how increase in fragment size leads to improved coverage

Techniques Used: Sequencing, Selection

Correlation between duplicate reads, spike-ins and the sequencing platforms.  a ,  b  Plots showing the frequency of distribution of duplicate reads for two cell line DNA samples during down sampling of the raw reads from HiSeq X Ten to the number of raw reads obtained from HiSeq 2500.  c  Box plot showing the difference in duplicate percentage when the same set of ten bisulphite libraries were spiked with 25% of 250 pM genomic library and 25% of 300 pM PhiX library
Figure Legend Snippet: Correlation between duplicate reads, spike-ins and the sequencing platforms. a , b Plots showing the frequency of distribution of duplicate reads for two cell line DNA samples during down sampling of the raw reads from HiSeq X Ten to the number of raw reads obtained from HiSeq 2500. c Box plot showing the difference in duplicate percentage when the same set of ten bisulphite libraries were spiked with 25% of 250 pM genomic library and 25% of 300 pM PhiX library

Techniques Used: Sequencing, Sampling

6) Product Images from "Unique, dual-indexed sequencing adapters with UMIs effectively eliminate index cross-talk and significantly improve sensitivity of massively parallel sequencing"

Article Title: Unique, dual-indexed sequencing adapters with UMIs effectively eliminate index cross-talk and significantly improve sensitivity of massively parallel sequencing

Journal: BMC Genomics

doi: 10.1186/s12864-017-4428-5

Level of cross-talk using unique, dual-matched indexed adapters on Illumina HiSeq 2500. The 96-well plate layout represents the adapter plate. A total of 35 adapters were synthesized. Seventeen human cell line libraries were prepared using IDT-synthesized unique, dual-matched indexed adapters (green), libraries were pooled, hybrid captured using a custom bait set, and then sequenced on a single lane of an Illumina HiSeq2500 flow cell. Numbers in each well represent the number of fragments that passed standard Illumina filters and demultiplexed using only perfect sequence matches on all TS-96 indices
Figure Legend Snippet: Level of cross-talk using unique, dual-matched indexed adapters on Illumina HiSeq 2500. The 96-well plate layout represents the adapter plate. A total of 35 adapters were synthesized. Seventeen human cell line libraries were prepared using IDT-synthesized unique, dual-matched indexed adapters (green), libraries were pooled, hybrid captured using a custom bait set, and then sequenced on a single lane of an Illumina HiSeq2500 flow cell. Numbers in each well represent the number of fragments that passed standard Illumina filters and demultiplexed using only perfect sequence matches on all TS-96 indices

Techniques Used: Synthesized, Flow Cytometry, Sequencing

7) Product Images from "Validation of multiple single nucleotide variation calls by additional exome analysis with a semiconductor sequencer to supplement data of whole-genome sequencing of a human population"

Article Title: Validation of multiple single nucleotide variation calls by additional exome analysis with a semiconductor sequencer to supplement data of whole-genome sequencing of a human population

Journal: BMC Genomics

doi: 10.1186/1471-2164-15-673

Box plots of read depths at concordant/discordant SNP loci among the NGSs and Omni 2.5-8. A) Box plots of the NGS read depth at the SNP loci showing concordance/discordance between the NGSs and Omni 2.5-8. The box indicates the first and third quartiles and the lines indicate the highest and lowest value that is within 1.5 x inter-quartile range. Outliers were omitted. The graph on the left indicates the distribution of read depths at HiSeq2500 SNP loci and the graph on the right indicates the distribution of read depths at Ion Proton and Omni 2.5-8 SNP loci. B) Box plots of the NGS read depths at the SNP loci showing concordance/discordance among the three platforms. The box indicates the first and third quartiles and the lines indicate the highest and lowest value that is within 1.5 x inter-quartile range. Outliers were omitted. The three columns (shaded gray rectangle) from right indicate the distribution of read depths of the discordant SNPs between the NGSs and Omni 2.5-8. The read depth at SNP loci that showed Ion Proton discordance against HiSeq 2500 and Omni 2.5-8 was significantly different. The read depths at SNP loci for SNPs that both HiSeq 2500 and Ion Proton failed to call are shown in the gray box.
Figure Legend Snippet: Box plots of read depths at concordant/discordant SNP loci among the NGSs and Omni 2.5-8. A) Box plots of the NGS read depth at the SNP loci showing concordance/discordance between the NGSs and Omni 2.5-8. The box indicates the first and third quartiles and the lines indicate the highest and lowest value that is within 1.5 x inter-quartile range. Outliers were omitted. The graph on the left indicates the distribution of read depths at HiSeq2500 SNP loci and the graph on the right indicates the distribution of read depths at Ion Proton and Omni 2.5-8 SNP loci. B) Box plots of the NGS read depths at the SNP loci showing concordance/discordance among the three platforms. The box indicates the first and third quartiles and the lines indicate the highest and lowest value that is within 1.5 x inter-quartile range. Outliers were omitted. The three columns (shaded gray rectangle) from right indicate the distribution of read depths of the discordant SNPs between the NGSs and Omni 2.5-8. The read depth at SNP loci that showed Ion Proton discordance against HiSeq 2500 and Omni 2.5-8 was significantly different. The read depths at SNP loci for SNPs that both HiSeq 2500 and Ion Proton failed to call are shown in the gray box.

Techniques Used: Next-Generation Sequencing

8) Product Images from "Guidelines for whole genome bisulphite sequencing of intact and FFPET DNA on the Illumina HiSeq X Ten"

Article Title: Guidelines for whole genome bisulphite sequencing of intact and FFPET DNA on the Illumina HiSeq X Ten

Journal: Epigenetics & Chromatin

doi: 10.1186/s13072-018-0194-0

Comparison of methylation correlation between HiSeq 2500 and HiSeq X Ten. a Correlation plots of methylation levels obtained from a cell line, a clinical sample and a FFPET sample sequenced on the Hiseq 2500 versus HiSeq X Ten (Pearson r > 0.94). b Correlation of methylation values obtained from HiSeq 2500 and HiSeq X Ten for a cell line, clinical sample and a FFPET sample after grouping them into four bins of methylation percentages. c Average kappa values for six sample pairs, including two cell lines, two clinical samples and two FFPE samples compared between the HiSeq 2500 and HiSeq X Ten platform. d Bar plot showing the distribution of percentage of discordant sites across the genome for a cell line, clinical sample and a FFPET sample
Figure Legend Snippet: Comparison of methylation correlation between HiSeq 2500 and HiSeq X Ten. a Correlation plots of methylation levels obtained from a cell line, a clinical sample and a FFPET sample sequenced on the Hiseq 2500 versus HiSeq X Ten (Pearson r > 0.94). b Correlation of methylation values obtained from HiSeq 2500 and HiSeq X Ten for a cell line, clinical sample and a FFPET sample after grouping them into four bins of methylation percentages. c Average kappa values for six sample pairs, including two cell lines, two clinical samples and two FFPE samples compared between the HiSeq 2500 and HiSeq X Ten platform. d Bar plot showing the distribution of percentage of discordant sites across the genome for a cell line, clinical sample and a FFPET sample

Techniques Used: Methylation, Formalin-fixed Paraffin-Embedded

Correlation between duplicate reads, spike-ins and the sequencing platforms.  a ,  b  Plots showing the frequency of distribution of duplicate reads for two cell line DNA samples during down sampling of the raw reads from HiSeq X Ten to the number of raw reads obtained from HiSeq 2500.  c  Box plot showing the difference in duplicate percentage when the same set of ten bisulphite libraries were spiked with 25% of 250 pM genomic library and 25% of 300 pM PhiX library
Figure Legend Snippet: Correlation between duplicate reads, spike-ins and the sequencing platforms. a , b Plots showing the frequency of distribution of duplicate reads for two cell line DNA samples during down sampling of the raw reads from HiSeq X Ten to the number of raw reads obtained from HiSeq 2500. c Box plot showing the difference in duplicate percentage when the same set of ten bisulphite libraries were spiked with 25% of 250 pM genomic library and 25% of 300 pM PhiX library

Techniques Used: Sequencing, Sampling

9) Product Images from "Unique, dual-indexed sequencing adapters with UMIs effectively eliminate index cross-talk and significantly improve sensitivity of massively parallel sequencing"

Article Title: Unique, dual-indexed sequencing adapters with UMIs effectively eliminate index cross-talk and significantly improve sensitivity of massively parallel sequencing

Journal: BMC Genomics

doi: 10.1186/s12864-017-4428-5

Level of cross-talk using unique, dual-matched indexed adapters on Illumina HiSeq 2500. The 96-well plate layout represents the adapter plate. A total of 35 adapters were synthesized. Seventeen human cell line libraries were prepared using IDT-synthesized unique, dual-matched indexed adapters (green), libraries were pooled, hybrid captured using a custom bait set, and then sequenced on a single lane of an Illumina HiSeq2500 flow cell. Numbers in each well represent the number of fragments that passed standard Illumina filters and demultiplexed using only perfect sequence matches on all TS-96 indices
Figure Legend Snippet: Level of cross-talk using unique, dual-matched indexed adapters on Illumina HiSeq 2500. The 96-well plate layout represents the adapter plate. A total of 35 adapters were synthesized. Seventeen human cell line libraries were prepared using IDT-synthesized unique, dual-matched indexed adapters (green), libraries were pooled, hybrid captured using a custom bait set, and then sequenced on a single lane of an Illumina HiSeq2500 flow cell. Numbers in each well represent the number of fragments that passed standard Illumina filters and demultiplexed using only perfect sequence matches on all TS-96 indices

Techniques Used: Synthesized, Flow Cytometry, Sequencing

10) Product Images from "Guidelines for whole genome bisulphite sequencing of intact and FFPET DNA on the Illumina HiSeq X Ten"

Article Title: Guidelines for whole genome bisulphite sequencing of intact and FFPET DNA on the Illumina HiSeq X Ten

Journal: Epigenetics & Chromatin

doi: 10.1186/s13072-018-0194-0

Coverage comparison between HiSeq 2500 and HiSeq X Ten. a Plot shows the fraction of genome covered at different depths for four samples sequenced together on one lane of the HiSeq 2500 versus when each of the samples is sequenced on a single lane of the HiSeq X Ten. The coverage plot for the HiSeq 2500 HO mode is the merged coverage obtained from multiplexing the four samples. b Plot shows the fraction of CpG sites covered at different depths when four clinical samples are sequenced together on one lane of the HiSeq 2500 versus when each of the samples is sequenced on a single lane of the HiSeq X Ten. c , d Box plot showing the coverage distribution across exons, intergenic regions, introns, promoter regions and repeat regions of the genome for a sample sequenced on one lane of HiSeq X Ten ( c ) and HiSeq 2500 ( d )
Figure Legend Snippet: Coverage comparison between HiSeq 2500 and HiSeq X Ten. a Plot shows the fraction of genome covered at different depths for four samples sequenced together on one lane of the HiSeq 2500 versus when each of the samples is sequenced on a single lane of the HiSeq X Ten. The coverage plot for the HiSeq 2500 HO mode is the merged coverage obtained from multiplexing the four samples. b Plot shows the fraction of CpG sites covered at different depths when four clinical samples are sequenced together on one lane of the HiSeq 2500 versus when each of the samples is sequenced on a single lane of the HiSeq X Ten. c , d Box plot showing the coverage distribution across exons, intergenic regions, introns, promoter regions and repeat regions of the genome for a sample sequenced on one lane of HiSeq X Ten ( c ) and HiSeq 2500 ( d )

Techniques Used: Multiplexing

Comparison of methylation correlation between HiSeq 2500 and HiSeq X Ten. a Correlation plots of methylation levels obtained from a cell line, a clinical sample and a FFPET sample sequenced on the Hiseq 2500 versus HiSeq X Ten (Pearson r > 0.94). b Correlation of methylation values obtained from HiSeq 2500 and HiSeq X Ten for a cell line, clinical sample and a FFPET sample after grouping them into four bins of methylation percentages. c Average kappa values for six sample pairs, including two cell lines, two clinical samples and two FFPE samples compared between the HiSeq 2500 and HiSeq X Ten platform. d Bar plot showing the distribution of percentage of discordant sites across the genome for a cell line, clinical sample and a FFPET sample
Figure Legend Snippet: Comparison of methylation correlation between HiSeq 2500 and HiSeq X Ten. a Correlation plots of methylation levels obtained from a cell line, a clinical sample and a FFPET sample sequenced on the Hiseq 2500 versus HiSeq X Ten (Pearson r > 0.94). b Correlation of methylation values obtained from HiSeq 2500 and HiSeq X Ten for a cell line, clinical sample and a FFPET sample after grouping them into four bins of methylation percentages. c Average kappa values for six sample pairs, including two cell lines, two clinical samples and two FFPE samples compared between the HiSeq 2500 and HiSeq X Ten platform. d Bar plot showing the distribution of percentage of discordant sites across the genome for a cell line, clinical sample and a FFPET sample

Techniques Used: Methylation, Formalin-fixed Paraffin-Embedded

Comparing different library preparation methods using genomic DNA on the HiSeq 2500.  a  Cluster plot of sequencing output metrics obtained from different library preparation methods from intact genomic DNA.  b  Bar graph showing fragment size distribution for different bead size selection for each library preparation method.  c  Plot with  x -axis indicating coverage and  y -axis indicating fragment size illustrating how increase in fragment size leads to improved coverage
Figure Legend Snippet: Comparing different library preparation methods using genomic DNA on the HiSeq 2500. a Cluster plot of sequencing output metrics obtained from different library preparation methods from intact genomic DNA. b Bar graph showing fragment size distribution for different bead size selection for each library preparation method. c Plot with x -axis indicating coverage and y -axis indicating fragment size illustrating how increase in fragment size leads to improved coverage

Techniques Used: Sequencing, Selection

Correlation between duplicate reads, spike-ins and the sequencing platforms.  a ,  b  Plots showing the frequency of distribution of duplicate reads for two cell line DNA samples during down sampling of the raw reads from HiSeq X Ten to the number of raw reads obtained from HiSeq 2500.  c  Box plot showing the difference in duplicate percentage when the same set of ten bisulphite libraries were spiked with 25% of 250 pM genomic library and 25% of 300 pM PhiX library
Figure Legend Snippet: Correlation between duplicate reads, spike-ins and the sequencing platforms. a , b Plots showing the frequency of distribution of duplicate reads for two cell line DNA samples during down sampling of the raw reads from HiSeq X Ten to the number of raw reads obtained from HiSeq 2500. c Box plot showing the difference in duplicate percentage when the same set of ten bisulphite libraries were spiked with 25% of 250 pM genomic library and 25% of 300 pM PhiX library

Techniques Used: Sequencing, Sampling

11) Product Images from "Guidelines for whole genome bisulphite sequencing of intact and FFPET DNA on the Illumina HiSeq X Ten"

Article Title: Guidelines for whole genome bisulphite sequencing of intact and FFPET DNA on the Illumina HiSeq X Ten

Journal: Epigenetics & Chromatin

doi: 10.1186/s13072-018-0194-0

Coverage comparison between HiSeq 2500 and HiSeq X Ten. a Plot shows the fraction of genome covered at different depths for four samples sequenced together on one lane of the HiSeq 2500 versus when each of the samples is sequenced on a single lane of the HiSeq X Ten. The coverage plot for the HiSeq 2500 HO mode is the merged coverage obtained from multiplexing the four samples. b Plot shows the fraction of CpG sites covered at different depths when four clinical samples are sequenced together on one lane of the HiSeq 2500 versus when each of the samples is sequenced on a single lane of the HiSeq X Ten. c , d Box plot showing the coverage distribution across exons, intergenic regions, introns, promoter regions and repeat regions of the genome for a sample sequenced on one lane of HiSeq X Ten ( c ) and HiSeq 2500 ( d )
Figure Legend Snippet: Coverage comparison between HiSeq 2500 and HiSeq X Ten. a Plot shows the fraction of genome covered at different depths for four samples sequenced together on one lane of the HiSeq 2500 versus when each of the samples is sequenced on a single lane of the HiSeq X Ten. The coverage plot for the HiSeq 2500 HO mode is the merged coverage obtained from multiplexing the four samples. b Plot shows the fraction of CpG sites covered at different depths when four clinical samples are sequenced together on one lane of the HiSeq 2500 versus when each of the samples is sequenced on a single lane of the HiSeq X Ten. c , d Box plot showing the coverage distribution across exons, intergenic regions, introns, promoter regions and repeat regions of the genome for a sample sequenced on one lane of HiSeq X Ten ( c ) and HiSeq 2500 ( d )

Techniques Used: Multiplexing

Comparison of methylation correlation between HiSeq 2500 and HiSeq X Ten. a Correlation plots of methylation levels obtained from a cell line, a clinical sample and a FFPET sample sequenced on the Hiseq 2500 versus HiSeq X Ten (Pearson r > 0.94). b Correlation of methylation values obtained from HiSeq 2500 and HiSeq X Ten for a cell line, clinical sample and a FFPET sample after grouping them into four bins of methylation percentages. c Average kappa values for six sample pairs, including two cell lines, two clinical samples and two FFPE samples compared between the HiSeq 2500 and HiSeq X Ten platform. d Bar plot showing the distribution of percentage of discordant sites across the genome for a cell line, clinical sample and a FFPET sample
Figure Legend Snippet: Comparison of methylation correlation between HiSeq 2500 and HiSeq X Ten. a Correlation plots of methylation levels obtained from a cell line, a clinical sample and a FFPET sample sequenced on the Hiseq 2500 versus HiSeq X Ten (Pearson r > 0.94). b Correlation of methylation values obtained from HiSeq 2500 and HiSeq X Ten for a cell line, clinical sample and a FFPET sample after grouping them into four bins of methylation percentages. c Average kappa values for six sample pairs, including two cell lines, two clinical samples and two FFPE samples compared between the HiSeq 2500 and HiSeq X Ten platform. d Bar plot showing the distribution of percentage of discordant sites across the genome for a cell line, clinical sample and a FFPET sample

Techniques Used: Methylation, Formalin-fixed Paraffin-Embedded

Comparing different library preparation methods using genomic DNA on the HiSeq 2500.  a  Cluster plot of sequencing output metrics obtained from different library preparation methods from intact genomic DNA.  b  Bar graph showing fragment size distribution for different bead size selection for each library preparation method.  c  Plot with  x -axis indicating coverage and  y -axis indicating fragment size illustrating how increase in fragment size leads to improved coverage
Figure Legend Snippet: Comparing different library preparation methods using genomic DNA on the HiSeq 2500. a Cluster plot of sequencing output metrics obtained from different library preparation methods from intact genomic DNA. b Bar graph showing fragment size distribution for different bead size selection for each library preparation method. c Plot with x -axis indicating coverage and y -axis indicating fragment size illustrating how increase in fragment size leads to improved coverage

Techniques Used: Sequencing, Selection

Correlation between duplicate reads, spike-ins and the sequencing platforms.  a ,  b  Plots showing the frequency of distribution of duplicate reads for two cell line DNA samples during down sampling of the raw reads from HiSeq X Ten to the number of raw reads obtained from HiSeq 2500.  c  Box plot showing the difference in duplicate percentage when the same set of ten bisulphite libraries were spiked with 25% of 250 pM genomic library and 25% of 300 pM PhiX library
Figure Legend Snippet: Correlation between duplicate reads, spike-ins and the sequencing platforms. a , b Plots showing the frequency of distribution of duplicate reads for two cell line DNA samples during down sampling of the raw reads from HiSeq X Ten to the number of raw reads obtained from HiSeq 2500. c Box plot showing the difference in duplicate percentage when the same set of ten bisulphite libraries were spiked with 25% of 250 pM genomic library and 25% of 300 pM PhiX library

Techniques Used: Sequencing, Sampling

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Amplification:

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Article Title: Novel mutational landscapes and expression signatures of lung squamous cell carcinoma
Article Snippet: The C1 Auto Prep System captures the dissociated single cells and the whole-transcriptome amplified cDNA was prepared on chip using the SMARTer Ultra Low RNA kit from Illumina (Clontech). .. Single-cell libraries were constructed with the use of the Illumina Nextera XT DNA Sample Preparation kit with 96 dual barcoded indices and were multiplexed and sequenced to a depth of 2–4 million reads (HiSeq 2500; Illumina) using 50-bp single-end sequencing.

Construct:

Article Title: Novel mutational landscapes and expression signatures of lung squamous cell carcinoma
Article Snippet: .. Single-cell libraries were constructed with the use of the Illumina Nextera XT DNA Sample Preparation kit with 96 dual barcoded indices and were multiplexed and sequenced to a depth of 2–4 million reads (HiSeq 2500; Illumina) using 50-bp single-end sequencing. .. The program RSEM [ ] was utilized to quantify transcript expression.

Single-cell Isolation:

Article Title: Novel mutational landscapes and expression signatures of lung squamous cell carcinoma
Article Snippet: Paragraph title: Single-cell isolation and RNA sequencing ... Single-cell libraries were constructed with the use of the Illumina Nextera XT DNA Sample Preparation kit with 96 dual barcoded indices and were multiplexed and sequenced to a depth of 2–4 million reads (HiSeq 2500; Illumina) using 50-bp single-end sequencing.

Expressing:

Article Title: RNA Sequencing Reveals a Slow to Fast Muscle Fiber Type Transition after Olanzapine Infusion in Rats
Article Snippet: Six libraries were pooled per HiSeq lane, followed by on-board cluster generation on a Rapid Run single-end flow cell and subsequent 50 cycles sequencing (v3 sequencing kit) according to the manufacturer's instructions (HiSeq 2500, Illumina). .. The uniquely mapped reads were used to calculate the normalized expression level of genes, as fragments per kilobase of exon per million fragments mapped (FPKM), using Cufflinks (v.2.2.1).

Article Title: Novel mutational landscapes and expression signatures of lung squamous cell carcinoma
Article Snippet: Single-cell libraries were constructed with the use of the Illumina Nextera XT DNA Sample Preparation kit with 96 dual barcoded indices and were multiplexed and sequenced to a depth of 2–4 million reads (HiSeq 2500; Illumina) using 50-bp single-end sequencing. .. The program RSEM [ ] was utilized to quantify transcript expression.

Article Title: A new synthetic toll-like receptor 1/2 ligand is an efficient adjuvant for peptide vaccination in a human volunteer
Article Snippet: Paragraph title: Transcriptome sequencing and analysis of differential gene expression* ... Single end sequencing was performed (HiSeq 2500; Illumina San Diego CA).

Real-time Polymerase Chain Reaction:

Article Title: Aligning to the sample-specific reference sequence to optimize the accuracy of next-generation sequencing analysis for hepatitis B virus
Article Snippet: The DNA library was quantified by Qubit fluorometer (Qubit dsDNA HS assay, Thermo Fisher Scientific, Life Technology) and real time PCR (KAPA Library Quantification Kit Illumina® platforms, KAPA biosystems). .. After it had been validated, the library was sequenced (HiSeq™ 2500; Illumina, San Diego, CA, USA).

Derivative Assay:

Article Title: Novel mutational landscapes and expression signatures of lung squamous cell carcinoma
Article Snippet: Single-cell libraries were constructed with the use of the Illumina Nextera XT DNA Sample Preparation kit with 96 dual barcoded indices and were multiplexed and sequenced to a depth of 2–4 million reads (HiSeq 2500; Illumina) using 50-bp single-end sequencing. .. Differential expression analysis was conducted on RSEM derived TPM (Transcripts Per Kilobase Million) values using the software AltAnalyzer [ ].

Flow Cytometry:

Article Title: RNA Sequencing Reveals a Slow to Fast Muscle Fiber Type Transition after Olanzapine Infusion in Rats
Article Snippet: .. Six libraries were pooled per HiSeq lane, followed by on-board cluster generation on a Rapid Run single-end flow cell and subsequent 50 cycles sequencing (v3 sequencing kit) according to the manufacturer's instructions (HiSeq 2500, Illumina). ..

Sequencing:

Article Title: Aligning to the sample-specific reference sequence to optimize the accuracy of next-generation sequencing analysis for hepatitis B virus
Article Snippet: Paragraph title: Viral genome sequencing using massively parallel NGS ... After it had been validated, the library was sequenced (HiSeq™ 2500; Illumina, San Diego, CA, USA).

Article Title: RNA Sequencing Reveals a Slow to Fast Muscle Fiber Type Transition after Olanzapine Infusion in Rats
Article Snippet: .. Six libraries were pooled per HiSeq lane, followed by on-board cluster generation on a Rapid Run single-end flow cell and subsequent 50 cycles sequencing (v3 sequencing kit) according to the manufacturer's instructions (HiSeq 2500, Illumina). ..

Article Title: Novel mutational landscapes and expression signatures of lung squamous cell carcinoma
Article Snippet: .. Single-cell libraries were constructed with the use of the Illumina Nextera XT DNA Sample Preparation kit with 96 dual barcoded indices and were multiplexed and sequenced to a depth of 2–4 million reads (HiSeq 2500; Illumina) using 50-bp single-end sequencing. .. The program RSEM [ ] was utilized to quantify transcript expression.

Article Title: A new synthetic toll-like receptor 1/2 ligand is an efficient adjuvant for peptide vaccination in a human volunteer
Article Snippet: .. Single end sequencing was performed (HiSeq 2500; Illumina San Diego CA). .. Mapping (hg19) (STAR software, V. 2.4.0), data processing and counts of mapped reads were computed (Cufflinks Tool Suite; Version 2.1.1).

Sonication:

Article Title: Aligning to the sample-specific reference sequence to optimize the accuracy of next-generation sequencing analysis for hepatitis B virus
Article Snippet: In brief, HBV DNA was fragmented using sonication and was cut into suitable sizes. .. After it had been validated, the library was sequenced (HiSeq™ 2500; Illumina, San Diego, CA, USA).

RNA Sequencing Assay:

Article Title: RNA Sequencing Reveals a Slow to Fast Muscle Fiber Type Transition after Olanzapine Infusion in Rats
Article Snippet: Paragraph title: RNA Sequencing ... Six libraries were pooled per HiSeq lane, followed by on-board cluster generation on a Rapid Run single-end flow cell and subsequent 50 cycles sequencing (v3 sequencing kit) according to the manufacturer's instructions (HiSeq 2500, Illumina).

Article Title: Novel mutational landscapes and expression signatures of lung squamous cell carcinoma
Article Snippet: Paragraph title: Single-cell isolation and RNA sequencing ... Single-cell libraries were constructed with the use of the Illumina Nextera XT DNA Sample Preparation kit with 96 dual barcoded indices and were multiplexed and sequenced to a depth of 2–4 million reads (HiSeq 2500; Illumina) using 50-bp single-end sequencing.

Isolation:

Article Title: RNA Sequencing Reveals a Slow to Fast Muscle Fiber Type Transition after Olanzapine Infusion in Rats
Article Snippet: Briefly, frozen muscle tissue was pulverized in liquid nitrogen using mortar and pestle, followed by bead mill homogenization (Bullet Blender, Next Advance) using stainless steel beads (Next Advance, cat# SSB14B) and mirVana RNA isolation kit (Life Technologies). .. Six libraries were pooled per HiSeq lane, followed by on-board cluster generation on a Rapid Run single-end flow cell and subsequent 50 cycles sequencing (v3 sequencing kit) according to the manufacturer's instructions (HiSeq 2500, Illumina).

Article Title: A new synthetic toll-like receptor 1/2 ligand is an efficient adjuvant for peptide vaccination in a human volunteer
Article Snippet: RNA was isolated from the granuloma center & margin and distal edge. .. Single end sequencing was performed (HiSeq 2500; Illumina San Diego CA).

Purification:

Article Title: Aligning to the sample-specific reference sequence to optimize the accuracy of next-generation sequencing analysis for hepatitis B virus
Article Snippet: These fragments were purified and were then end-repaired and A-tailed using DNA Polymerase I Klenow Fragment (3′ → 5′ exo-) (New England Biolabs, Ipswich, MA, USA). .. After it had been validated, the library was sequenced (HiSeq™ 2500; Illumina, San Diego, CA, USA).

Polymerase Chain Reaction:

Article Title: Aligning to the sample-specific reference sequence to optimize the accuracy of next-generation sequencing analysis for hepatitis B virus
Article Snippet: DNA fragments ligated with indexed adapters were amplified using 10–18 cycles of PCR reaction. .. After it had been validated, the library was sequenced (HiSeq™ 2500; Illumina, San Diego, CA, USA).

Chromatin Immunoprecipitation:

Article Title: Novel mutational landscapes and expression signatures of lung squamous cell carcinoma
Article Snippet: The C1 Auto Prep System captures the dissociated single cells and the whole-transcriptome amplified cDNA was prepared on chip using the SMARTer Ultra Low RNA kit from Illumina (Clontech). .. Single-cell libraries were constructed with the use of the Illumina Nextera XT DNA Sample Preparation kit with 96 dual barcoded indices and were multiplexed and sequenced to a depth of 2–4 million reads (HiSeq 2500; Illumina) using 50-bp single-end sequencing.

Software:

Article Title: Novel mutational landscapes and expression signatures of lung squamous cell carcinoma
Article Snippet: Single-cell libraries were constructed with the use of the Illumina Nextera XT DNA Sample Preparation kit with 96 dual barcoded indices and were multiplexed and sequenced to a depth of 2–4 million reads (HiSeq 2500; Illumina) using 50-bp single-end sequencing. .. Differential expression analysis was conducted on RSEM derived TPM (Transcripts Per Kilobase Million) values using the software AltAnalyzer [ ].

Article Title: A new synthetic toll-like receptor 1/2 ligand is an efficient adjuvant for peptide vaccination in a human volunteer
Article Snippet: Single end sequencing was performed (HiSeq 2500; Illumina San Diego CA). .. Mapping (hg19) (STAR software, V. 2.4.0), data processing and counts of mapped reads were computed (Cufflinks Tool Suite; Version 2.1.1).

Electrophoresis:

Article Title: Aligning to the sample-specific reference sequence to optimize the accuracy of next-generation sequencing analysis for hepatitis B virus
Article Snippet: Experion Automated Electrophoresis System (Bio-Rad Laboratories, Hercules, CA, USA) was used to validate the size of the library. .. After it had been validated, the library was sequenced (HiSeq™ 2500; Illumina, San Diego, CA, USA).

Sample Prep:

Article Title: RNA Sequencing Reveals a Slow to Fast Muscle Fiber Type Transition after Olanzapine Infusion in Rats
Article Snippet: The cDNA libraries were prepared using the TruSeq RNA Sample Prep Kit v2 (Illumina) as per the manufacturer’s instructions. .. Six libraries were pooled per HiSeq lane, followed by on-board cluster generation on a Rapid Run single-end flow cell and subsequent 50 cycles sequencing (v3 sequencing kit) according to the manufacturer's instructions (HiSeq 2500, Illumina).

Article Title: Novel mutational landscapes and expression signatures of lung squamous cell carcinoma
Article Snippet: .. Single-cell libraries were constructed with the use of the Illumina Nextera XT DNA Sample Preparation kit with 96 dual barcoded indices and were multiplexed and sequenced to a depth of 2–4 million reads (HiSeq 2500; Illumina) using 50-bp single-end sequencing. .. The program RSEM [ ] was utilized to quantify transcript expression.

Homogenization:

Article Title: RNA Sequencing Reveals a Slow to Fast Muscle Fiber Type Transition after Olanzapine Infusion in Rats
Article Snippet: Briefly, frozen muscle tissue was pulverized in liquid nitrogen using mortar and pestle, followed by bead mill homogenization (Bullet Blender, Next Advance) using stainless steel beads (Next Advance, cat# SSB14B) and mirVana RNA isolation kit (Life Technologies). .. Six libraries were pooled per HiSeq lane, followed by on-board cluster generation on a Rapid Run single-end flow cell and subsequent 50 cycles sequencing (v3 sequencing kit) according to the manufacturer's instructions (HiSeq 2500, Illumina).

Next-Generation Sequencing:

Article Title: Aligning to the sample-specific reference sequence to optimize the accuracy of next-generation sequencing analysis for hepatitis B virus
Article Snippet: Paragraph title: Viral genome sequencing using massively parallel NGS ... After it had been validated, the library was sequenced (HiSeq™ 2500; Illumina, San Diego, CA, USA).

Concentration Assay:

Article Title: RNA Sequencing Reveals a Slow to Fast Muscle Fiber Type Transition after Olanzapine Infusion in Rats
Article Snippet: The final product was assessed for its size distribution using Bioanalyzer DNA High Sensitivity Kit (Agilent Technologies) and for its concentration using Kapa library quantification kit (Kapa Biosystems). .. Six libraries were pooled per HiSeq lane, followed by on-board cluster generation on a Rapid Run single-end flow cell and subsequent 50 cycles sequencing (v3 sequencing kit) according to the manufacturer's instructions (HiSeq 2500, Illumina).

Lysis:

Article Title: Novel mutational landscapes and expression signatures of lung squamous cell carcinoma
Article Snippet: For single-cell analysis cell capture, lysis, reverse transcription, and cDNA amplification were performed on the C1 integrated fluidic circuit (IFC) for mRNA-seq on a Fluidigm C1 Single-Cell Auto Prep System following the manufacturer’s protocol (Fluidigm Corporation, CA). .. Single-cell libraries were constructed with the use of the Illumina Nextera XT DNA Sample Preparation kit with 96 dual barcoded indices and were multiplexed and sequenced to a depth of 2–4 million reads (HiSeq 2500; Illumina) using 50-bp single-end sequencing.

Single-cell Analysis:

Article Title: Novel mutational landscapes and expression signatures of lung squamous cell carcinoma
Article Snippet: For single-cell analysis cell capture, lysis, reverse transcription, and cDNA amplification were performed on the C1 integrated fluidic circuit (IFC) for mRNA-seq on a Fluidigm C1 Single-Cell Auto Prep System following the manufacturer’s protocol (Fluidigm Corporation, CA). .. Single-cell libraries were constructed with the use of the Illumina Nextera XT DNA Sample Preparation kit with 96 dual barcoded indices and were multiplexed and sequenced to a depth of 2–4 million reads (HiSeq 2500; Illumina) using 50-bp single-end sequencing.

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  • 99
    Illumina Inc hiseq 2500 sequencing system
    Illustrated representation of the optimized barcoded cDNA library preparation procedure. Total RNA from formalin-fixed paraffin-embedded (FFPE) specimens ( n = 18), are ligated with 3′ adapters ( n = 18), purified on a 15% PAGE gel (using size markers (SM) RNA oligonucleotides) and excised after SYBR ® gold staining of the gel. The combined purified products are ligated with a 5′ adapter, run on a 12% PAGE gel, excised and purified. The dual-adapter ligated small-RNA products are reverse transcribed, subjected to a pilot PCR for identification of adequate amplification cycle. A large-scale PCR is performed, and the products are separated on a 2% agarose gel, where the upper band (cDNA library) is purified and subjected to sequencing on an Illumina <t>HiSeq-2500</t> sequencer.
    Hiseq 2500 Sequencing System, supplied by Illumina Inc, used in various techniques. Bioz Stars score: 99/100, based on 234 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/hiseq 2500 sequencing system/product/Illumina Inc
    Average 99 stars, based on 234 article reviews
    Price from $9.99 to $1999.99
    hiseq 2500 sequencing system - by Bioz Stars, 2020-04
    99/100 stars
      Buy from Supplier

    99
    Illumina Inc hiseq 2500
    Zinc-related and gli cluster genes were upregulated during phagocytosis. ( A ) Total RNA was extracted from the macrophage phagocytic cells infected with conidia of A. fumigatus , and libraries were prepared from 2 µg of total RNA. High-throughput sequencing was performed as paired-end 100 bp sequencing using a <t>HiSeq</t> 2500 (Illumina, Inc., USA). mRNA-seq reads were mapped using the TopHat software tool to obtain an alignment file. ( B ) Total RNA was extracted from the phagocytic cells infected with conidia after 0 and 1 h postinfection, and cDNA was synthesized using a TOPscript™ cDNA synthesis kit (Enzynomics). Then, qRT-PCR was performed using KAPA SYBR ® for LightCycler ® 480 with the primer sets for the indicated genes. ( C ) The transcriptomic profile during phagocytosis is shown. The gene expression profile was grouped into zinc, copper, iron, and gliotoxin metabolism. Upregulated genes were found in all categories, and ZafA and GliZ were upregulated during phagocytosis. Up- and downregulated genes are colored to show fold change. * p
    Hiseq 2500, supplied by Illumina Inc, used in various techniques. Bioz Stars score: 99/100, based on 690 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/hiseq 2500/product/Illumina Inc
    Average 99 stars, based on 690 article reviews
    Price from $9.99 to $1999.99
    hiseq 2500 - by Bioz Stars, 2020-04
    99/100 stars
      Buy from Supplier

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    Illustrated representation of the optimized barcoded cDNA library preparation procedure. Total RNA from formalin-fixed paraffin-embedded (FFPE) specimens ( n = 18), are ligated with 3′ adapters ( n = 18), purified on a 15% PAGE gel (using size markers (SM) RNA oligonucleotides) and excised after SYBR ® gold staining of the gel. The combined purified products are ligated with a 5′ adapter, run on a 12% PAGE gel, excised and purified. The dual-adapter ligated small-RNA products are reverse transcribed, subjected to a pilot PCR for identification of adequate amplification cycle. A large-scale PCR is performed, and the products are separated on a 2% agarose gel, where the upper band (cDNA library) is purified and subjected to sequencing on an Illumina HiSeq-2500 sequencer.

    Journal: International Journal of Molecular Sciences

    Article Title: Evaluation and Adaptation of a Laboratory-Based cDNA Library Preparation Protocol for Retrospective Sequencing of Archived MicroRNAs from up to 35-Year-Old Clinical FFPE Specimens

    doi: 10.3390/ijms18030627

    Figure Lengend Snippet: Illustrated representation of the optimized barcoded cDNA library preparation procedure. Total RNA from formalin-fixed paraffin-embedded (FFPE) specimens ( n = 18), are ligated with 3′ adapters ( n = 18), purified on a 15% PAGE gel (using size markers (SM) RNA oligonucleotides) and excised after SYBR ® gold staining of the gel. The combined purified products are ligated with a 5′ adapter, run on a 12% PAGE gel, excised and purified. The dual-adapter ligated small-RNA products are reverse transcribed, subjected to a pilot PCR for identification of adequate amplification cycle. A large-scale PCR is performed, and the products are separated on a 2% agarose gel, where the upper band (cDNA library) is purified and subjected to sequencing on an Illumina HiSeq-2500 sequencer.

    Article Snippet: The cDNA libraries were then sequenced (single-read 50 cycles) on a HiSeq 2500 Sequencing System (SY-401-2501, Illumina, San Diego, CA, USA).

    Techniques: cDNA Library Assay, Formalin-fixed Paraffin-Embedded, Purification, Polyacrylamide Gel Electrophoresis, Staining, Polymerase Chain Reaction, Amplification, Agarose Gel Electrophoresis, Sequencing

    Zinc-related and gli cluster genes were upregulated during phagocytosis. ( A ) Total RNA was extracted from the macrophage phagocytic cells infected with conidia of A. fumigatus , and libraries were prepared from 2 µg of total RNA. High-throughput sequencing was performed as paired-end 100 bp sequencing using a HiSeq 2500 (Illumina, Inc., USA). mRNA-seq reads were mapped using the TopHat software tool to obtain an alignment file. ( B ) Total RNA was extracted from the phagocytic cells infected with conidia after 0 and 1 h postinfection, and cDNA was synthesized using a TOPscript™ cDNA synthesis kit (Enzynomics). Then, qRT-PCR was performed using KAPA SYBR ® for LightCycler ® 480 with the primer sets for the indicated genes. ( C ) The transcriptomic profile during phagocytosis is shown. The gene expression profile was grouped into zinc, copper, iron, and gliotoxin metabolism. Upregulated genes were found in all categories, and ZafA and GliZ were upregulated during phagocytosis. Up- and downregulated genes are colored to show fold change. * p

    Journal: International Journal of Molecular Sciences

    Article Title: The Role of Zinc in Gliotoxin Biosynthesis of Aspergillus fumigatus

    doi: 10.3390/ijms20246192

    Figure Lengend Snippet: Zinc-related and gli cluster genes were upregulated during phagocytosis. ( A ) Total RNA was extracted from the macrophage phagocytic cells infected with conidia of A. fumigatus , and libraries were prepared from 2 µg of total RNA. High-throughput sequencing was performed as paired-end 100 bp sequencing using a HiSeq 2500 (Illumina, Inc., USA). mRNA-seq reads were mapped using the TopHat software tool to obtain an alignment file. ( B ) Total RNA was extracted from the phagocytic cells infected with conidia after 0 and 1 h postinfection, and cDNA was synthesized using a TOPscript™ cDNA synthesis kit (Enzynomics). Then, qRT-PCR was performed using KAPA SYBR ® for LightCycler ® 480 with the primer sets for the indicated genes. ( C ) The transcriptomic profile during phagocytosis is shown. The gene expression profile was grouped into zinc, copper, iron, and gliotoxin metabolism. Upregulated genes were found in all categories, and ZafA and GliZ were upregulated during phagocytosis. Up- and downregulated genes are colored to show fold change. * p

    Article Snippet: High-throughput sequencing was performed as paired-end 100 bp sequencing using a HiSeq 2500 (Illumina, Inc., California, USA). mRNA-seq reads were mapped using the TopHat software tool to obtain an alignment file.

    Techniques: Infection, Next-Generation Sequencing, Sequencing, Software, Synthesized, Quantitative RT-PCR, Expressing

    Diagram of the validation procedure of the noninvasive chromosome screening (NICS) assay. Briefly, D3 embryos achieved via intracytoplasmic sperm injection (ICSI) were placed in blastocyst culture medium. Both the D3–D5 culture medium and the corresponding whole embryos were collected and used for WGA by multiple annealing and looping-based amplification cycles (MALBAC). Whole-genome–amplified products from both the D3–D5 culture medium and the embryo were sequenced using an Illumina HiSeq 2500 platform. The chromosome ploidy information was obtained from both the culture medium and the corresponding embryo.

    Journal: Proceedings of the National Academy of Sciences of the United States of America

    Article Title: Noninvasive chromosome screening of human embryos by genome sequencing of embryo culture medium for in vitro fertilization

    doi: 10.1073/pnas.1613294113

    Figure Lengend Snippet: Diagram of the validation procedure of the noninvasive chromosome screening (NICS) assay. Briefly, D3 embryos achieved via intracytoplasmic sperm injection (ICSI) were placed in blastocyst culture medium. Both the D3–D5 culture medium and the corresponding whole embryos were collected and used for WGA by multiple annealing and looping-based amplification cycles (MALBAC). Whole-genome–amplified products from both the D3–D5 culture medium and the embryo were sequenced using an Illumina HiSeq 2500 platform. The chromosome ploidy information was obtained from both the culture medium and the corresponding embryo.

    Article Snippet: The MALBAC single-cell WGA method was used to amplify DNA from the culture medium, as well as from the embryos, following the manufacturer’s protocol (catalog no. YK001B; Yikon Genomics), and used to construct the sequencing libraries (NEBNext Ultra DNA Kit; New England Biolabs) for sequencing on an Illumina HiSeq 2500 platform, yielding ∼2 million sequencing reads on each sample.

    Techniques: Injection, Whole Genome Amplification, Multiple Annealing and Looping–Based Amplification Cycles, Amplification

    Experimental scheme. ( A ) After differentiation, the cell cultures were dissociated to single-cell suspensions and loaded onto a Fluidigm C1 Single-Cell Auto Prep Array for mRNA-Seq. On the arrays, the cells were lysed, and reverse transcription of the mRNA and PCR amplification of the cDNA were performed using the C1 Single-Cell Auto Prep System (Fluidigm). Libraries were then prepared using the Nextera XT DNA Library Prep Kit and mosquito HTS liquid handler (TTP). For the final PCR reactions, we used a Bio-Rad 384 Thermal Cycler. Libraries were pooled and sequenced on an Illumina HiSeq 2500. ( B ) WA09 human embryonic stem cells were differentiated in vitro to the pancreatic lineage. Cells from stages 1 and 2 were collected and analyzed using the procedures outlined in A . Two independent cells from stage 1 (cell A and cell B) and two cells from stage 2 (cell C and cell D) were selected and yielded with similar cDNA concentrations (mean [SD] concentration = 0.38 [0.04] ng/µL). For library preparation, we tested 2-µL, 4-µL, and 8-µL final volume reactions, with four replicates per reaction volume.

    Journal: Jala (Charlottesville, Va.)

    Article Title: Miniaturization Technologies for Efficient Single-Cell Library Preparation for Next-Generation Sequencing

    doi: 10.1177/2211068216630741

    Figure Lengend Snippet: Experimental scheme. ( A ) After differentiation, the cell cultures were dissociated to single-cell suspensions and loaded onto a Fluidigm C1 Single-Cell Auto Prep Array for mRNA-Seq. On the arrays, the cells were lysed, and reverse transcription of the mRNA and PCR amplification of the cDNA were performed using the C1 Single-Cell Auto Prep System (Fluidigm). Libraries were then prepared using the Nextera XT DNA Library Prep Kit and mosquito HTS liquid handler (TTP). For the final PCR reactions, we used a Bio-Rad 384 Thermal Cycler. Libraries were pooled and sequenced on an Illumina HiSeq 2500. ( B ) WA09 human embryonic stem cells were differentiated in vitro to the pancreatic lineage. Cells from stages 1 and 2 were collected and analyzed using the procedures outlined in A . Two independent cells from stage 1 (cell A and cell B) and two cells from stage 2 (cell C and cell D) were selected and yielded with similar cDNA concentrations (mean [SD] concentration = 0.38 [0.04] ng/µL). For library preparation, we tested 2-µL, 4-µL, and 8-µL final volume reactions, with four replicates per reaction volume.

    Article Snippet: Sequencing The 48 pooled libraries were sequenced on one lane of an Illumina HiSeq 2500 at an average total read depth of 5.6 million reads per sample (ST3).

    Techniques: Polymerase Chain Reaction, Amplification, In Vitro, Concentration Assay