Benchmarking of the Oxford Nanopore MinION sequencing for quantitative and qualitative assessment of cDNA populations

To assess the performance of the Oxford Nanopore Technologies MinION sequencing platform, cDNAs from the External RNA Controls Consortium (ERCC) RNA Spike-In mix were sequenced. This mix mimics mammalian mRNA species and consists of 92 polyadenylated transcripts with known concentration. cDNA librar...

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Published in:	Scientific reports Vol. 6; no. 1; p. 31602
Main Authors:	Oikonomopoulos, Spyros, Wang, Yu Chang, Djambazian, Haig, Badescu, Dunarel, Ragoussis, Jiannis
Format:	Journal Article
Language:	English
Published:	London Nature Publishing Group UK 24.08.2016 Nature Publishing Group
Subjects:	38/23 38/39 38/91 45 45/23 45/91 631/1647/2017/2003 631/1647/2217/2018 631/208/514/1949 631/208/514/2254 Agreements Animals DNA, Complementary - genetics Gene expression Gene Library HEK293 Cells High-Throughput Nucleotide Sequencing - methods Humanities and Social Sciences Humans Isoforms multidisciplinary Nanopores Performance assessment Performance evaluation Polyadenylation Population Science
ISSN:	2045-2322, 2045-2322
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Abstract	To assess the performance of the Oxford Nanopore Technologies MinION sequencing platform, cDNAs from the External RNA Controls Consortium (ERCC) RNA Spike-In mix were sequenced. This mix mimics mammalian mRNA species and consists of 92 polyadenylated transcripts with known concentration. cDNA libraries were generated using a template switching protocol to facilitate the direct comparison between different sequencing platforms. The MinION performance was assessed for its ability to sequence the cDNAs directly with good accuracy in terms of abundance and full length. The abundance of the ERCC cDNA molecules sequenced by MinION agreed with their expected concentration. No length or GC content bias was observed. The majority of cDNAs were sequenced as full length. Additionally, a complex cDNA population derived from a human HEK-293 cell line was sequenced on an Illumina HiSeq 2500, PacBio RS II and ONT MinION platforms. We observed that there was a good agreement in the measured cDNA abundance between PacBio RS II and ONT MinION (r pearson = 0.82, isoforms with length more than 700bp) and between Illumina HiSeq 2500 and ONT MinION (r pearson = 0.75). This indicates that the ONT MinION can sequence quantitatively both long and short full length cDNA molecules.
AbstractList	To assess the performance of the Oxford Nanopore Technologies MinION sequencing platform, cDNAs from the External RNA Controls Consortium (ERCC) RNA Spike-In mix were sequenced. This mix mimics mammalian mRNA species and consists of 92 polyadenylated transcripts with known concentration. cDNA libraries were generated using a template switching protocol to facilitate the direct comparison between different sequencing platforms. The MinION performance was assessed for its ability to sequence the cDNAs directly with good accuracy in terms of abundance and full length. The abundance of the ERCC cDNA molecules sequenced by MinION agreed with their expected concentration. No length or GC content bias was observed. The majority of cDNAs were sequenced as full length. Additionally, a complex cDNA population derived from a human HEK-293 cell line was sequenced on an Illumina HiSeq 2500, PacBio RS II and ONT MinION platforms. We observed that there was a good agreement in the measured cDNA abundance between PacBio RS II and ONT MinION (rpearson = 0.82, isoforms with length more than 700bp) and between Illumina HiSeq 2500 and ONT MinION (rpearson = 0.75). This indicates that the ONT MinION can sequence quantitatively both long and short full length cDNA molecules. To assess the performance of the Oxford Nanopore Technologies MinION sequencing platform, cDNAs from the External RNA Controls Consortium (ERCC) RNA Spike-In mix were sequenced. This mix mimics mammalian mRNA species and consists of 92 polyadenylated transcripts with known concentration. cDNA libraries were generated using a template switching protocol to facilitate the direct comparison between different sequencing platforms. The MinION performance was assessed for its ability to sequence the cDNAs directly with good accuracy in terms of abundance and full length. The abundance of the ERCC cDNA molecules sequenced by MinION agreed with their expected concentration. No length or GC content bias was observed. The majority of cDNAs were sequenced as full length. Additionally, a complex cDNA population derived from a human HEK-293 cell line was sequenced on an Illumina HiSeq 2500, PacBio RS II and ONT MinION platforms. We observed that there was a good agreement in the measured cDNA abundance between PacBio RS II and ONT MinION (r pearson = 0.82, isoforms with length more than 700bp) and between Illumina HiSeq 2500 and ONT MinION (r pearson = 0.75). This indicates that the ONT MinION can sequence quantitatively both long and short full length cDNA molecules. To assess the performance of the Oxford Nanopore Technologies MinION sequencing platform, cDNAs from the External RNA Controls Consortium (ERCC) RNA Spike-In mix were sequenced. This mix mimics mammalian mRNA species and consists of 92 polyadenylated transcripts with known concentration. cDNA libraries were generated using a template switching protocol to facilitate the direct comparison between different sequencing platforms. The MinION performance was assessed for its ability to sequence the cDNAs directly with good accuracy in terms of abundance and full length. The abundance of the ERCC cDNA molecules sequenced by MinION agreed with their expected concentration. No length or GC content bias was observed. The majority of cDNAs were sequenced as full length. Additionally, a complex cDNA population derived from a human HEK-293 cell line was sequenced on an Illumina HiSeq 2500, PacBio RS II and ONT MinION platforms. We observed that there was a good agreement in the measured cDNA abundance between PacBio RS II and ONT MinION (rpearson = 0.82, isoforms with length more than 700bp) and between Illumina HiSeq 2500 and ONT MinION (rpearson = 0.75). This indicates that the ONT MinION can sequence quantitatively both long and short full length cDNA molecules.To assess the performance of the Oxford Nanopore Technologies MinION sequencing platform, cDNAs from the External RNA Controls Consortium (ERCC) RNA Spike-In mix were sequenced. This mix mimics mammalian mRNA species and consists of 92 polyadenylated transcripts with known concentration. cDNA libraries were generated using a template switching protocol to facilitate the direct comparison between different sequencing platforms. The MinION performance was assessed for its ability to sequence the cDNAs directly with good accuracy in terms of abundance and full length. The abundance of the ERCC cDNA molecules sequenced by MinION agreed with their expected concentration. No length or GC content bias was observed. The majority of cDNAs were sequenced as full length. Additionally, a complex cDNA population derived from a human HEK-293 cell line was sequenced on an Illumina HiSeq 2500, PacBio RS II and ONT MinION platforms. We observed that there was a good agreement in the measured cDNA abundance between PacBio RS II and ONT MinION (rpearson = 0.82, isoforms with length more than 700bp) and between Illumina HiSeq 2500 and ONT MinION (rpearson = 0.75). This indicates that the ONT MinION can sequence quantitatively both long and short full length cDNA molecules.
ArticleNumber	31602
Author	Wang, Yu Chang Djambazian, Haig Badescu, Dunarel Ragoussis, Jiannis Oikonomopoulos, Spyros
Author_xml	– sequence: 1 givenname: Spyros surname: Oikonomopoulos fullname: Oikonomopoulos, Spyros organization: Department of Human Genetics, McGill University and Genome Quebec Innovation Centre, 740 Dr. Penfield Avenue, McGill University – sequence: 2 givenname: Yu Chang surname: Wang fullname: Wang, Yu Chang organization: Department of Human Genetics, McGill University and Genome Quebec Innovation Centre, 740 Dr. Penfield Avenue, McGill University – sequence: 3 givenname: Haig surname: Djambazian fullname: Djambazian, Haig organization: Department of Human Genetics, McGill University and Genome Quebec Innovation Centre, 740 Dr. Penfield Avenue, McGill University – sequence: 4 givenname: Dunarel surname: Badescu fullname: Badescu, Dunarel organization: Department of Human Genetics, McGill University and Genome Quebec Innovation Centre, 740 Dr. Penfield Avenue, McGill University – sequence: 5 givenname: Jiannis surname: Ragoussis fullname: Ragoussis, Jiannis organization: Department of Human Genetics, McGill University and Genome Quebec Innovation Centre, 740 Dr. Penfield Avenue, McGill University, Department of Bioengineering, McGill University, Department of Biochemistry, Center of Innovation in Personalized Medicine, Cancer and Mutagen Unit, King Fahd Center for Medical Research, King Abdulaziz University
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Snippet	To assess the performance of the Oxford Nanopore Technologies MinION sequencing platform, cDNAs from the External RNA Controls Consortium (ERCC) RNA Spike-In...
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SubjectTerms	38/23 38/39 38/91 45 45/23 45/91 631/1647/2017/2003 631/1647/2217/2018 631/208/514/1949 631/208/514/2254 Agreements Animals DNA, Complementary - genetics Gene expression Gene Library HEK293 Cells High-Throughput Nucleotide Sequencing - methods Humanities and Social Sciences Humans Isoforms multidisciplinary Nanopores Performance assessment Performance evaluation Polyadenylation Population Science
Title	Benchmarking of the Oxford Nanopore MinION sequencing for quantitative and qualitative assessment of cDNA populations
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