Amplification of four genes of influenza A viruses using a degenerate primer set in a one step RT-PCR method

Naresh Jindal; Yogesh Chander; Martha de Abin; Srinand Sreevatsan; David Stallknecht; David A. Halvorson; Sagar M. Goyal

doi:10.1016/j.jviromet.2009.05.003

Amplification of four genes of influenza A viruses using a degenerate primer set in a one step RT-PCR method

Naresh Jindal, Yogesh Chander, Martha de Abin, Srinand Sreevatsan, David Stallknecht, David A. Halvorson, Sagar M. Goyal

Research output: Contribution to journal › Article › peer-review

13 Scopus citations

Abstract

We designed a degenerate primer set that yielded full-length amplification of hemagglutinin (HA), neuraminidase (NA), matrix (M), and non-structural protein (NSP) genes of influenza A viruses in a single reaction mixture. These four genes were amplified from 15 HA (1-15) and 9 NA (1-9) subtypes of influenza A viruses of avian (n = 16) origin. In addition, 272 field isolates of avian origin were tested by this method. Full-length amplification of HA, NA, M, and NSP genes was obtained in 242 (88.9%), 254 (93.4%), 268 (98.5%), and 268 (98.5%) isolates, respectively. No gene was amplified in four isolates. Of these four isolates, two were subtyped as H4N6, one as H7N7, and one as H10N7. Amplification was successful for all 4 genes of H1N1, H2N3, and H3N2 isolates of swine influenza. Also, all four genes were amplified in one equine influenza (H3N8) isolate and seven isolates of human origin (H1N1 and H3N2). This appears to be the first study using degenerate primer set for full-length amplification of four genes of influenza A viruses in a single reaction. Further studies are needed to determine if this primer set can be used for subtyping of influenza virus isolates.

Original language	English (US)
Pages (from-to)	163-166
Number of pages	4
Journal	Journal of Virological Methods
Volume	160
Issue number	1-2
DOIs	https://doi.org/10.1016/j.jviromet.2009.05.003
State	Published - Sep 2009

Bibliographical note

Funding Information:
This work has been funded in whole or in part with federal funds from the National Institute of Allergy and Infectious Diseases, National Institutes of Health, Department of Health and Human Services, under Contract No. HHSN266200700007C. Its contents are solely the responsibility of the authors and do not necessarily represent the official views of the NIH. We thank Brundaban Panigrahy and Chinta Lamichhane for providing avian influenza viruses and Michael Osterholm for helpful comments.

Keywords

Degenerate primers
Gene amplification
Influenza A virus
Reverse transcription-polymerase chain reaction (RT-PCR)

UN SDGs

This output contributes to the following UN Sustainable Development Goals (SDGs)

Publisher link

10.1016/j.jviromet.2009.05.003

Find It

Find It at U of M Libraries

Cite this

@article{469504dd9e024a96b140291522f6ef7d,

title = "Amplification of four genes of influenza A viruses using a degenerate primer set in a one step RT-PCR method",

abstract = "We designed a degenerate primer set that yielded full-length amplification of hemagglutinin (HA), neuraminidase (NA), matrix (M), and non-structural protein (NSP) genes of influenza A viruses in a single reaction mixture. These four genes were amplified from 15 HA (1-15) and 9 NA (1-9) subtypes of influenza A viruses of avian (n = 16) origin. In addition, 272 field isolates of avian origin were tested by this method. Full-length amplification of HA, NA, M, and NSP genes was obtained in 242 (88.9%), 254 (93.4%), 268 (98.5%), and 268 (98.5%) isolates, respectively. No gene was amplified in four isolates. Of these four isolates, two were subtyped as H4N6, one as H7N7, and one as H10N7. Amplification was successful for all 4 genes of H1N1, H2N3, and H3N2 isolates of swine influenza. Also, all four genes were amplified in one equine influenza (H3N8) isolate and seven isolates of human origin (H1N1 and H3N2). This appears to be the first study using degenerate primer set for full-length amplification of four genes of influenza A viruses in a single reaction. Further studies are needed to determine if this primer set can be used for subtyping of influenza virus isolates.",

keywords = "Degenerate primers, Gene amplification, Influenza A virus, Reverse transcription-polymerase chain reaction (RT-PCR)",

author = "Naresh Jindal and Yogesh Chander and {de Abin}, Martha and Srinand Sreevatsan and David Stallknecht and Halvorson, {David A.} and Goyal, {Sagar M.}",

note = "Funding Information: This work has been funded in whole or in part with federal funds from the National Institute of Allergy and Infectious Diseases, National Institutes of Health, Department of Health and Human Services, under Contract No. HHSN266200700007C. Its contents are solely the responsibility of the authors and do not necessarily represent the official views of the NIH. We thank Brundaban Panigrahy and Chinta Lamichhane for providing avian influenza viruses and Michael Osterholm for helpful comments.",

year = "2009",

month = sep,

doi = "10.1016/j.jviromet.2009.05.003",

language = "English (US)",

volume = "160",

pages = "163--166",

journal = "Journal of Virological Methods",

issn = "0166-0934",

publisher = "Elsevier",

number = "1-2",

}

TY - JOUR

T1 - Amplification of four genes of influenza A viruses using a degenerate primer set in a one step RT-PCR method

AU - Jindal, Naresh

AU - Chander, Yogesh

AU - de Abin, Martha

AU - Sreevatsan, Srinand

AU - Stallknecht, David

AU - Halvorson, David A.

AU - Goyal, Sagar M.

N1 - Funding Information: This work has been funded in whole or in part with federal funds from the National Institute of Allergy and Infectious Diseases, National Institutes of Health, Department of Health and Human Services, under Contract No. HHSN266200700007C. Its contents are solely the responsibility of the authors and do not necessarily represent the official views of the NIH. We thank Brundaban Panigrahy and Chinta Lamichhane for providing avian influenza viruses and Michael Osterholm for helpful comments.

PY - 2009/9

Y1 - 2009/9

N2 - We designed a degenerate primer set that yielded full-length amplification of hemagglutinin (HA), neuraminidase (NA), matrix (M), and non-structural protein (NSP) genes of influenza A viruses in a single reaction mixture. These four genes were amplified from 15 HA (1-15) and 9 NA (1-9) subtypes of influenza A viruses of avian (n = 16) origin. In addition, 272 field isolates of avian origin were tested by this method. Full-length amplification of HA, NA, M, and NSP genes was obtained in 242 (88.9%), 254 (93.4%), 268 (98.5%), and 268 (98.5%) isolates, respectively. No gene was amplified in four isolates. Of these four isolates, two were subtyped as H4N6, one as H7N7, and one as H10N7. Amplification was successful for all 4 genes of H1N1, H2N3, and H3N2 isolates of swine influenza. Also, all four genes were amplified in one equine influenza (H3N8) isolate and seven isolates of human origin (H1N1 and H3N2). This appears to be the first study using degenerate primer set for full-length amplification of four genes of influenza A viruses in a single reaction. Further studies are needed to determine if this primer set can be used for subtyping of influenza virus isolates.

AB - We designed a degenerate primer set that yielded full-length amplification of hemagglutinin (HA), neuraminidase (NA), matrix (M), and non-structural protein (NSP) genes of influenza A viruses in a single reaction mixture. These four genes were amplified from 15 HA (1-15) and 9 NA (1-9) subtypes of influenza A viruses of avian (n = 16) origin. In addition, 272 field isolates of avian origin were tested by this method. Full-length amplification of HA, NA, M, and NSP genes was obtained in 242 (88.9%), 254 (93.4%), 268 (98.5%), and 268 (98.5%) isolates, respectively. No gene was amplified in four isolates. Of these four isolates, two were subtyped as H4N6, one as H7N7, and one as H10N7. Amplification was successful for all 4 genes of H1N1, H2N3, and H3N2 isolates of swine influenza. Also, all four genes were amplified in one equine influenza (H3N8) isolate and seven isolates of human origin (H1N1 and H3N2). This appears to be the first study using degenerate primer set for full-length amplification of four genes of influenza A viruses in a single reaction. Further studies are needed to determine if this primer set can be used for subtyping of influenza virus isolates.

KW - Degenerate primers

KW - Gene amplification

KW - Influenza A virus

KW - Reverse transcription-polymerase chain reaction (RT-PCR)

UR - http://www.scopus.com/inward/record.url?scp=67549100777&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=67549100777&partnerID=8YFLogxK

U2 - 10.1016/j.jviromet.2009.05.003

DO - 10.1016/j.jviromet.2009.05.003

M3 - Article

C2 - 19447141

AN - SCOPUS:67549100777

SN - 0166-0934

VL - 160

SP - 163

EP - 166

JO - Journal of Virological Methods

JF - Journal of Virological Methods

IS - 1-2

ER -

Amplification of four genes of influenza A viruses using a degenerate primer set in a one step RT-PCR method

Abstract

Bibliographical note

Keywords

UN SDGs

Publisher link

Find It

Other files and links

Fingerprint

Cite this