An Amplicon-Based Approach for the Whole-Genome Sequencing of Human Metapneumovirus
Abstract
:1. Introduction
2. Materials and Methods
2.1. Primer Design
2.2. Clinical Specimens and HMPV Isolation
2.3. Reverse Transcription
2.4. HMPV Genome Amplification
2.5. Library Preparation and Sequencing
2.6. Viral Assembly
2.7. Phylogenetic Analysis
3. Results and Discussion
3.1. Designing Primers to Amplify the HMPV Genome
3.2. HMPV Genome RT-PCR Performance
3.3. Genome Sequencing, Assembly and Analysis
4. Conclusions
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Acknowledgments
Conflicts of Interest
References
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Assay | Primer Name | Sequence (5’-3’) | Position (nt) * | PCR Amplicon Size (bp) |
---|---|---|---|---|
PCR1 | HMPV1_F | GGGACAAATAAAAATGTCTCTTCA | 41 | 4125 |
HMPV1_R | CTTCCTGTGCTRACYTTRCA | 4165 | ||
PCR2 | HMPV2_F | ACAGCAGCRGGRATYAATGT | 4074 | 4010 |
HMPV2_R | TAGTACTGAAYTGAGCATGYTCAG | 8083 | ||
PCR3 | HMPV3_F | AACTGTTAACATGGAAAGATGTGATG | 7823 | 3229 |
HMPV3_R | TAAGCTGGAACWGAWGCTG | 11,051 | ||
PCR4 | HMPV4_F | TCAATAGGGAGTCTRTGTCARGAA | 9730 | 3675 |
HMPV4_R | GRCAAAAAAACCGTATACATYC | 13,404 |
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Tulloch, R.L.; Kok, J.; Carter, I.; Dwyer, D.E.; Eden, J.-S. An Amplicon-Based Approach for the Whole-Genome Sequencing of Human Metapneumovirus. Viruses 2021, 13, 499. https://doi.org/10.3390/v13030499
Tulloch RL, Kok J, Carter I, Dwyer DE, Eden J-S. An Amplicon-Based Approach for the Whole-Genome Sequencing of Human Metapneumovirus. Viruses. 2021; 13(3):499. https://doi.org/10.3390/v13030499
Chicago/Turabian StyleTulloch, Rachel L., Jen Kok, Ian Carter, Dominic E. Dwyer, and John-Sebastian Eden. 2021. "An Amplicon-Based Approach for the Whole-Genome Sequencing of Human Metapneumovirus" Viruses 13, no. 3: 499. https://doi.org/10.3390/v13030499