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Using 454 technology for long-PCR based sequencing of the complete mitochondrial genome from single Haemonchus contortus (Nematoda)

BACKGROUND: Mitochondrial (mt) genomes represent a rich source of molecular markers for a range of applications, including population genetics, systematics, epidemiology and ecology. In the present study, we used 454 technology (or the GS20, massively parallel picolitre reactor platform) to determin...

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Autores principales: Jex, Aaron R, Hu, Min, Littlewood, D Timothy J, Waeschenbach, Andrea, Gasser, Robin B
Formato: Texto
Lenguaje:English
Publicado: BioMed Central 2008
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2254599/
https://www.ncbi.nlm.nih.gov/pubmed/18190685
http://dx.doi.org/10.1186/1471-2164-9-11
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author Jex, Aaron R
Hu, Min
Littlewood, D Timothy J
Waeschenbach, Andrea
Gasser, Robin B
author_facet Jex, Aaron R
Hu, Min
Littlewood, D Timothy J
Waeschenbach, Andrea
Gasser, Robin B
author_sort Jex, Aaron R
collection PubMed
description BACKGROUND: Mitochondrial (mt) genomes represent a rich source of molecular markers for a range of applications, including population genetics, systematics, epidemiology and ecology. In the present study, we used 454 technology (or the GS20, massively parallel picolitre reactor platform) to determine the complete mt genome of Haemonchus contortus (Nematoda: Trichostrongylidae), a parasite of substantial agricultural, veterinary and economic significance. We validate this approach by comparison with mt sequences from publicly available expressed sequence tag (EST) and genomic survey sequence (GSS) data sets. RESULTS: The complete mt genome of Haemonchus contortus was sequenced directly from long-PCR amplified template utilizing genomic DNA (~20–40 ng) from a single adult male using 454 technology. A single contig was assembled and compared against mt sequences mined from publicly available EST (NemBLAST) and GSS datasets. The comparison demonstrated that the 454 technology platform is reliable for the sequencing of AT-rich mt genomes from nematodes. The mt genome sequenced for Haemonchus contortus was 14,055 bp in length and was highly AT-rich (78.1%). In accordance with other chromadorean nematodes studied to date, the mt genome of H. contortus contained 36 genes (12 protein coding, 22 tRNAs, rrnL and rrnS) and was similar in structure, size and gene arrangement to those characterized previously for members of the Strongylida. CONCLUSION: The present study demonstrates the utility of 454 technology for the rapid determination of mt genome sequences from tiny amounts of DNA and reveals a wealth of mt genomic data in current databases available for mining. This approach provides a novel platform for high-throughput sequencing of mt genomes from nematodes and other organisms.
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spelling pubmed-22545992008-02-27 Using 454 technology for long-PCR based sequencing of the complete mitochondrial genome from single Haemonchus contortus (Nematoda) Jex, Aaron R Hu, Min Littlewood, D Timothy J Waeschenbach, Andrea Gasser, Robin B BMC Genomics Research Article BACKGROUND: Mitochondrial (mt) genomes represent a rich source of molecular markers for a range of applications, including population genetics, systematics, epidemiology and ecology. In the present study, we used 454 technology (or the GS20, massively parallel picolitre reactor platform) to determine the complete mt genome of Haemonchus contortus (Nematoda: Trichostrongylidae), a parasite of substantial agricultural, veterinary and economic significance. We validate this approach by comparison with mt sequences from publicly available expressed sequence tag (EST) and genomic survey sequence (GSS) data sets. RESULTS: The complete mt genome of Haemonchus contortus was sequenced directly from long-PCR amplified template utilizing genomic DNA (~20–40 ng) from a single adult male using 454 technology. A single contig was assembled and compared against mt sequences mined from publicly available EST (NemBLAST) and GSS datasets. The comparison demonstrated that the 454 technology platform is reliable for the sequencing of AT-rich mt genomes from nematodes. The mt genome sequenced for Haemonchus contortus was 14,055 bp in length and was highly AT-rich (78.1%). In accordance with other chromadorean nematodes studied to date, the mt genome of H. contortus contained 36 genes (12 protein coding, 22 tRNAs, rrnL and rrnS) and was similar in structure, size and gene arrangement to those characterized previously for members of the Strongylida. CONCLUSION: The present study demonstrates the utility of 454 technology for the rapid determination of mt genome sequences from tiny amounts of DNA and reveals a wealth of mt genomic data in current databases available for mining. This approach provides a novel platform for high-throughput sequencing of mt genomes from nematodes and other organisms. BioMed Central 2008-01-11 /pmc/articles/PMC2254599/ /pubmed/18190685 http://dx.doi.org/10.1186/1471-2164-9-11 Text en Copyright © 2008 Jex et al; licensee BioMed Central Ltd. http://creativecommons.org/licenses/by/2.0 This is an Open Access article distributed under the terms of the Creative Commons Attribution License ( (http://creativecommons.org/licenses/by/2.0) ), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
spellingShingle Research Article
Jex, Aaron R
Hu, Min
Littlewood, D Timothy J
Waeschenbach, Andrea
Gasser, Robin B
Using 454 technology for long-PCR based sequencing of the complete mitochondrial genome from single Haemonchus contortus (Nematoda)
title Using 454 technology for long-PCR based sequencing of the complete mitochondrial genome from single Haemonchus contortus (Nematoda)
title_full Using 454 technology for long-PCR based sequencing of the complete mitochondrial genome from single Haemonchus contortus (Nematoda)
title_fullStr Using 454 technology for long-PCR based sequencing of the complete mitochondrial genome from single Haemonchus contortus (Nematoda)
title_full_unstemmed Using 454 technology for long-PCR based sequencing of the complete mitochondrial genome from single Haemonchus contortus (Nematoda)
title_short Using 454 technology for long-PCR based sequencing of the complete mitochondrial genome from single Haemonchus contortus (Nematoda)
title_sort using 454 technology for long-pcr based sequencing of the complete mitochondrial genome from single haemonchus contortus (nematoda)
topic Research Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2254599/
https://www.ncbi.nlm.nih.gov/pubmed/18190685
http://dx.doi.org/10.1186/1471-2164-9-11
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