Selection and validation of suitable reference genes for qPCR gene expression analysis in goats and sheep under Peste des petits ruminants virus (PPRV), lineage IV infection
Identifieur interne : 000695 ( Main/Exploration ); précédent : 000694; suivant : 000696Selection and validation of suitable reference genes for qPCR gene expression analysis in goats and sheep under Peste des petits ruminants virus (PPRV), lineage IV infection
Auteurs : Amit Ranjan Sahu [Inde] ; Sajad Ahmad Wani [États-Unis] ; Shikha Saxena ; Kaushal Kishor Rajak ; Dheeraj Chaudhary [Inde] ; Aditya Prasad Sahoo [Inde] ; Alok Khanduri ; Aruna Pandey ; Piyali Mondal ; Waseem Akram Malla ; Raja Ishaq Nabi Khan ; Ashok Kumar Tiwari ; Bina Mishra ; D. Muthuchelvan [Inde] ; Bishnu Prasad Mishra ; Raj Kumar Singh ; Ravi Kumar Gandham [Inde]Source :
- Scientific Reports [ 2045-2322 ] ; 2018.
Abstract
Identification of suitable candidate reference genes is an important prerequisite for validating the gene expression data obtained from downstream analysis of RNA sequencing using quantitative real time PCR (qRT-PCR). Though existence of a universal reference gene is myth, commonly used reference genes can be assessed for expression stability to confer their suitability to be used as candidate reference genes in gene expression studies. In this study, we evaluated the expression stability of ten most commonly used reference genes (
Url:
DOI: 10.1038/s41598-018-34236-7
PubMed: 30374051
PubMed Central: 6206032
Affiliations:
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expression analysis in goats and sheep under <italic>Peste des petits ruminants virus (PPRV)</italic>, lineage IV infection</title><author><name sortKey="Sahu, Amit Ranjan" sort="Sahu, Amit Ranjan" uniqKey="Sahu A" first="Amit Ranjan" last="Sahu">Amit Ranjan Sahu</name><affiliation><nlm:aff id="Aff1">Division of Veterinary Biotechnology, ICAR-IVRI, Izatnagar, Bareilly, UP 243122 India</nlm:aff><wicri:noCountry code="subfield">UP 243122 India</wicri:noCountry></affiliation><affiliation wicri:level="1"><nlm:aff id="Aff2">DBT-National Institute of Animal Biotechnology, Hyderabad, 500075 India</nlm:aff><country xml:lang="fr">Inde</country><wicri:regionArea>DBT-National Institute of Animal Biotechnology, Hyderabad</wicri:regionArea><wicri:noRegion>Hyderabad</wicri:noRegion></affiliation></author><author><name sortKey="Wani, Sajad Ahmad" sort="Wani, Sajad Ahmad" uniqKey="Wani S" first="Sajad Ahmad" last="Wani">Sajad Ahmad Wani</name><affiliation><nlm:aff id="Aff1">Division of Veterinary Biotechnology, ICAR-IVRI, Izatnagar, Bareilly, UP 243122 India</nlm:aff><wicri:noCountry code="subfield">UP 243122 India</wicri:noCountry></affiliation><affiliation wicri:level="2"><nlm:aff id="Aff3"><institution-wrap><institution-id institution-id-type="ISNI">0000 0001 2285 7943</institution-id><institution-id institution-id-type="GRID">grid.261331.4</institution-id><institution>The Ohio State University,</institution></institution-wrap>Columbus, Ohio 43210 USA</nlm:aff><country xml:lang="fr">États-Unis</country><placeName><region type="state">Ohio</region></placeName><wicri:cityArea>Columbus</wicri:cityArea></affiliation></author><author><name sortKey="Saxena, Shikha" sort="Saxena, Shikha" uniqKey="Saxena S" first="Shikha" last="Saxena">Shikha Saxena</name><affiliation><nlm:aff id="Aff1">Division of Veterinary Biotechnology, ICAR-IVRI, Izatnagar, Bareilly, UP 243122 India</nlm:aff><wicri:noCountry code="subfield">UP 243122 India</wicri:noCountry></affiliation></author><author><name sortKey="Rajak, Kaushal Kishor" sort="Rajak, Kaushal Kishor" uniqKey="Rajak K" first="Kaushal Kishor" last="Rajak">Kaushal Kishor Rajak</name><affiliation><nlm:aff id="Aff4">Division of Biological Products, ICAR-IVRI, Izatnagar, Bareilly, UP 243122 India</nlm:aff><wicri:noCountry code="subfield">UP 243122 India</wicri:noCountry></affiliation></author><author><name sortKey="Chaudhary, Dheeraj" sort="Chaudhary, Dheeraj" uniqKey="Chaudhary D" first="Dheeraj" last="Chaudhary">Dheeraj Chaudhary</name><affiliation wicri:level="1"><nlm:aff id="Aff5">Division of Virology, ICAR-IVRI, Mukteshwar Campus, Nainital, 263138 India</nlm:aff><country xml:lang="fr">Inde</country><wicri:regionArea>Division of Virology, ICAR-IVRI, Mukteshwar Campus, Nainital</wicri:regionArea><wicri:noRegion>Nainital</wicri:noRegion></affiliation></author><author><name sortKey="Sahoo, Aditya Prasad" sort="Sahoo, Aditya Prasad" uniqKey="Sahoo A" first="Aditya Prasad" last="Sahoo">Aditya Prasad Sahoo</name><affiliation wicri:level="1"><nlm:aff id="Aff6">ICAR-Directorate of Foot and Mouth Disease, Mukteshwar, Nainital, 263138 India</nlm:aff><country xml:lang="fr">Inde</country><wicri:regionArea>ICAR-Directorate of Foot and Mouth Disease, Mukteshwar, Nainital</wicri:regionArea><wicri:noRegion>Nainital</wicri:noRegion></affiliation></author><author><name sortKey="Khanduri, Alok" sort="Khanduri, Alok" uniqKey="Khanduri A" first="Alok" last="Khanduri">Alok Khanduri</name><affiliation><nlm:aff id="Aff1">Division of Veterinary Biotechnology, ICAR-IVRI, Izatnagar, Bareilly, UP 243122 India</nlm:aff><wicri:noCountry code="subfield">UP 243122 India</wicri:noCountry></affiliation></author><author><name sortKey="Pandey, Aruna" sort="Pandey, Aruna" uniqKey="Pandey A" first="Aruna" last="Pandey">Aruna Pandey</name><affiliation><nlm:aff id="Aff1">Division of Veterinary Biotechnology, ICAR-IVRI, Izatnagar, Bareilly, UP 243122 India</nlm:aff><wicri:noCountry code="subfield">UP 243122 India</wicri:noCountry></affiliation></author><author><name sortKey="Mondal, Piyali" sort="Mondal, Piyali" uniqKey="Mondal P" first="Piyali" last="Mondal">Piyali Mondal</name><affiliation><nlm:aff id="Aff1">Division of Veterinary Biotechnology, ICAR-IVRI, Izatnagar, Bareilly, UP 243122 India</nlm:aff><wicri:noCountry code="subfield">UP 243122 India</wicri:noCountry></affiliation></author><author><name sortKey="Malla, Waseem Akram" sort="Malla, Waseem Akram" uniqKey="Malla W" first="Waseem Akram" last="Malla">Waseem Akram Malla</name><affiliation><nlm:aff id="Aff1">Division of Veterinary Biotechnology, ICAR-IVRI, Izatnagar, Bareilly, UP 243122 India</nlm:aff><wicri:noCountry code="subfield">UP 243122 India</wicri:noCountry></affiliation></author><author><name sortKey="Khan, Raja Ishaq Nabi" sort="Khan, Raja Ishaq Nabi" uniqKey="Khan R" first="Raja Ishaq Nabi" last="Khan">Raja Ishaq Nabi Khan</name><affiliation><nlm:aff id="Aff1">Division of Veterinary Biotechnology, ICAR-IVRI, Izatnagar, Bareilly, UP 243122 India</nlm:aff><wicri:noCountry code="subfield">UP 243122 India</wicri:noCountry></affiliation></author><author><name sortKey="Tiwari, Ashok Kumar" sort="Tiwari, Ashok Kumar" uniqKey="Tiwari A" first="Ashok Kumar" last="Tiwari">Ashok Kumar Tiwari</name><affiliation><nlm:aff id="Aff7">Division of Biological Standardization, ICAR-IVRI, Izatnagar, Bareilly, UP 243122 India</nlm:aff><wicri:noCountry code="subfield">UP 243122 India</wicri:noCountry></affiliation></author><author><name sortKey="Mishra, Bina" sort="Mishra, Bina" uniqKey="Mishra B" first="Bina" last="Mishra">Bina Mishra</name><affiliation><nlm:aff id="Aff4">Division of Biological Products, ICAR-IVRI, Izatnagar, Bareilly, UP 243122 India</nlm:aff><wicri:noCountry code="subfield">UP 243122 India</wicri:noCountry></affiliation></author><author><name sortKey="Muthuchelvan, D" sort="Muthuchelvan, D" uniqKey="Muthuchelvan D" first="D." last="Muthuchelvan">D. Muthuchelvan</name><affiliation wicri:level="1"><nlm:aff id="Aff5">Division of Virology, ICAR-IVRI, Mukteshwar Campus, Nainital, 263138 India</nlm:aff><country xml:lang="fr">Inde</country><wicri:regionArea>Division of Virology, ICAR-IVRI, Mukteshwar Campus, Nainital</wicri:regionArea><wicri:noRegion>Nainital</wicri:noRegion></affiliation></author><author><name sortKey="Mishra, Bishnu Prasad" sort="Mishra, Bishnu Prasad" uniqKey="Mishra B" first="Bishnu Prasad" last="Mishra">Bishnu Prasad Mishra</name><affiliation><nlm:aff id="Aff1">Division of Veterinary Biotechnology, ICAR-IVRI, Izatnagar, Bareilly, UP 243122 India</nlm:aff><wicri:noCountry code="subfield">UP 243122 India</wicri:noCountry></affiliation></author><author><name sortKey="Singh, Raj Kumar" sort="Singh, Raj Kumar" uniqKey="Singh R" first="Raj Kumar" last="Singh">Raj Kumar Singh</name><affiliation><nlm:aff id="Aff1">Division of Veterinary Biotechnology, ICAR-IVRI, Izatnagar, Bareilly, UP 243122 India</nlm:aff><wicri:noCountry code="subfield">UP 243122 India</wicri:noCountry></affiliation></author><author><name sortKey="Gandham, Ravi Kumar" sort="Gandham, Ravi Kumar" uniqKey="Gandham R" first="Ravi Kumar" last="Gandham">Ravi Kumar Gandham</name><affiliation><nlm:aff id="Aff1">Division of Veterinary Biotechnology, ICAR-IVRI, Izatnagar, Bareilly, UP 243122 India</nlm:aff><wicri:noCountry code="subfield">UP 243122 India</wicri:noCountry></affiliation><affiliation wicri:level="1"><nlm:aff id="Aff2">DBT-National Institute of Animal Biotechnology, Hyderabad, 500075 India</nlm:aff><country xml:lang="fr">Inde</country><wicri:regionArea>DBT-National Institute of Animal Biotechnology, Hyderabad</wicri:regionArea><wicri:noRegion>Hyderabad</wicri:noRegion></affiliation></author></analytic><series><title level="j">Scientific Reports</title><idno type="eISSN">2045-2322</idno><imprint><date when="2018">2018</date></imprint></series></biblStruct></sourceDesc></fileDesc><profileDesc><textClass></textClass></profileDesc></teiHeader><front><div type="abstract" xml:lang="en"><p id="Par1">Identification of suitable candidate reference genes is an important prerequisite for validating the gene expression data obtained from downstream analysis of RNA sequencing using quantitative real time PCR (qRT-PCR). Though existence of a universal reference gene is myth, commonly used reference genes can be assessed for expression stability to confer their suitability to be used as candidate reference genes in gene expression studies. In this study, we evaluated the expression stability of ten most commonly used reference genes (<italic>GAPDH</italic>, <italic>ACTB</italic>, <italic>HSP90</italic>, <italic>HMBS</italic>, <italic>18S rRNA</italic>, <italic>B2M</italic>, <italic>POLR2A</italic>, <italic>HPRT1</italic>, <italic>ACAC</italic>, <italic>YWHAZ</italic>) in fourteen different Peste des petits ruminants virus (PPRV) infected tissues of goats and sheep. RefFinder and RankAggreg software were used to deduce comprehensive ranking of reference genes. Our results suggested <italic>HMBS</italic> and <italic>B2M</italic> in goats and <italic>HMBS</italic> and <italic>HPRT1</italic> in sheep can be used as suitable endogenous controls in gene expression studies of PPRV infection irrespective of tissues and condition as a whole, thus eliminating the use of tissue specific/ condition specific endogenous controls. We report for the first time suitable reference genes for gene expression studies in PPRV infected tissues. The reference genes determined here can be useful for future studies on gene expression in sheep and goat infected with PPRV, thus saving extra efforts and time of repeating the reference gene determination and validation.</p></div></front><back><div1 type="bibliography"><listBibl><biblStruct><analytic><author><name sortKey="Fang, Z" uniqKey="Fang Z">Z Fang</name></author><author><name sortKey="Cui, X" uniqKey="Cui X">X Cui</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Marioni, Jc" uniqKey="Marioni J">JC Marioni</name></author><author><name sortKey="Mason, Ce" uniqKey="Mason C">CE Mason</name></author><author><name sortKey="Mane, Sm" uniqKey="Mane S">SM Mane</name></author><author><name sortKey="Stephens, M" uniqKey="Stephens M">M Stephens</name></author><author><name sortKey="Gilad, Y" uniqKey="Gilad Y">Y Gilad</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Pickrell, Jk" uniqKey="Pickrell J">JK 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uniqKey="Mishra B" first="Bishnu Prasad" last="Mishra">Bishnu Prasad Mishra</name><name sortKey="Mondal, Piyali" sort="Mondal, Piyali" uniqKey="Mondal P" first="Piyali" last="Mondal">Piyali Mondal</name><name sortKey="Pandey, Aruna" sort="Pandey, Aruna" uniqKey="Pandey A" first="Aruna" last="Pandey">Aruna Pandey</name><name sortKey="Rajak, Kaushal Kishor" sort="Rajak, Kaushal Kishor" uniqKey="Rajak K" first="Kaushal Kishor" last="Rajak">Kaushal Kishor Rajak</name><name sortKey="Saxena, Shikha" sort="Saxena, Shikha" uniqKey="Saxena S" first="Shikha" last="Saxena">Shikha Saxena</name><name sortKey="Singh, Raj Kumar" sort="Singh, Raj Kumar" uniqKey="Singh R" first="Raj Kumar" last="Singh">Raj Kumar Singh</name><name sortKey="Tiwari, Ashok Kumar" sort="Tiwari, Ashok Kumar" uniqKey="Tiwari A" first="Ashok Kumar" last="Tiwari">Ashok Kumar Tiwari</name></noCountry><country name="Inde"><noRegion><name sortKey="Sahu, Amit Ranjan" sort="Sahu, Amit Ranjan" uniqKey="Sahu A" first="Amit Ranjan" last="Sahu">Amit Ranjan Sahu</name></noRegion><name sortKey="Chaudhary, Dheeraj" sort="Chaudhary, Dheeraj" uniqKey="Chaudhary D" first="Dheeraj" last="Chaudhary">Dheeraj Chaudhary</name><name sortKey="Gandham, Ravi Kumar" sort="Gandham, Ravi Kumar" uniqKey="Gandham R" first="Ravi Kumar" last="Gandham">Ravi Kumar Gandham</name><name sortKey="Muthuchelvan, D" sort="Muthuchelvan, D" uniqKey="Muthuchelvan D" first="D." last="Muthuchelvan">D. Muthuchelvan</name><name sortKey="Sahoo, Aditya Prasad" sort="Sahoo, Aditya Prasad" uniqKey="Sahoo A" first="Aditya Prasad" last="Sahoo">Aditya Prasad Sahoo</name></country><country name="États-Unis"><region name="Ohio"><name sortKey="Wani, Sajad Ahmad" sort="Wani, Sajad Ahmad" uniqKey="Wani S" first="Sajad Ahmad" last="Wani">Sajad Ahmad Wani</name></region></country></tree></affiliations></record>Pour manipuler ce document sous Unix (Dilib)
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