Identification of Reference Genes for Quantitative Real-Time PCR in Date Palm (Phoenix dactylifera L.) Subjected to Drought and Salinity
Date palm is an important crop plant in the arid and semi-arid regions supporting human population in the Middle East and North Africa. These areas have been largely affected by drought and salinity due to insufficient rainfall and improper irrigation practices. Date palm is a relatively salt- and d...
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| Veröffentlicht in: | PloS one 2016-11, Vol.11 (11), p.e0166216-e0166216 |
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| creator | V Patankar, Himanshu M Assaha, Dekoum V Al-Yahyai, Rashid Sunkar, Ramanjulu Yaish, Mahmoud W |
| description | Date palm is an important crop plant in the arid and semi-arid regions supporting human population in the Middle East and North Africa. These areas have been largely affected by drought and salinity due to insufficient rainfall and improper irrigation practices. Date palm is a relatively salt- and drought-tolerant plant and more recently efforts have been directed to identifying genes and pathways that confer stress tolerance in this species. Quantitative real-time PCR (qPCR) is a promising technique for the analysis of stress-induced differential gene expression, which involves the use of stable reference genes for normalizing gene expression. In an attempt to find the best reference genes for date palm's drought and salinity research, we evaluated the stability of 12 most commonly used reference genes using the geNorm, NormFinder, BestKeeper statistical algorithms and the comparative ΔCT method. The comprehensive results revealed that HEAT SHOCK PROTEIN (HSP), UBIQUITIN (UBQ) and YTH domain-containing family protein (YT521) were stable in drought-stressed leaves whereas GLYCERALDEHYDE-3-PHOSPHATE DEHYDROGENASE (GAPDH), ACTIN and TUBULIN were stable in drought-stressed roots. On the other hand, SMALL SUBUNIT RIBOSOMAL RNA (25S), YT521 and 18S ribosomal RNA (18S); and UBQ, ACTIN and ELONGATION FACTOR 1-ALPHA (eEF1a) were stable in leaves and roots, respectively, under salt stress. The stability of these reference genes was verified by using the abiotic stress-responsive CYTOSOLIC Cu/Zn SUPEROXIDE DISMUTASE (Cyt-Cu/Zn SOD), an ABA RECEPTOR, and a PROLINE TRANSPORTER 2 (PRO) genes. A combination of top 2 or 3 stable reference genes were found to be suitable for normalization of the target gene expression and will facilitate gene expression analysis studies aimed at identifying functional genes associated with drought and salinity tolerance in date palm. |
| doi_str_mv | 10.1371/journal.pone.0166216 |
| format | Article |
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Date palm is a relatively salt- and drought-tolerant plant and more recently efforts have been directed to identifying genes and pathways that confer stress tolerance in this species. Quantitative real-time PCR (qPCR) is a promising technique for the analysis of stress-induced differential gene expression, which involves the use of stable reference genes for normalizing gene expression. In an attempt to find the best reference genes for date palm's drought and salinity research, we evaluated the stability of 12 most commonly used reference genes using the geNorm, NormFinder, BestKeeper statistical algorithms and the comparative ΔCT method. The comprehensive results revealed that HEAT SHOCK PROTEIN (HSP), UBIQUITIN (UBQ) and YTH domain-containing family protein (YT521) were stable in drought-stressed leaves whereas GLYCERALDEHYDE-3-PHOSPHATE DEHYDROGENASE (GAPDH), ACTIN and TUBULIN were stable in drought-stressed roots. On the other hand, SMALL SUBUNIT RIBOSOMAL RNA (25S), YT521 and 18S ribosomal RNA (18S); and UBQ, ACTIN and ELONGATION FACTOR 1-ALPHA (eEF1a) were stable in leaves and roots, respectively, under salt stress. The stability of these reference genes was verified by using the abiotic stress-responsive CYTOSOLIC Cu/Zn SUPEROXIDE DISMUTASE (Cyt-Cu/Zn SOD), an ABA RECEPTOR, and a PROLINE TRANSPORTER 2 (PRO) genes. A combination of top 2 or 3 stable reference genes were found to be suitable for normalization of the target gene expression and will facilitate gene expression analysis studies aimed at identifying functional genes associated with drought and salinity tolerance in date palm.</description><identifier>ISSN: 1932-6203</identifier><identifier>EISSN: 1932-6203</identifier><identifier>DOI: 10.1371/journal.pone.0166216</identifier><identifier>PMID: 27824922</identifier><language>eng</language><publisher>United States: Public Library of Science</publisher><subject>Abiotic stress ; Actin ; Africa, Northern ; Algorithms ; Analysis ; Arid regions ; Arid zones ; Biology and Life Sciences ; Copper ; Drought ; Droughts ; Ecology and Environmental Sciences ; Elongation ; Gene expression ; Gene Expression Profiling - methods ; Gene Expression Regulation, Plant - genetics ; Genes ; Genes, Plant - genetics ; Genomes ; Glyceraldehyde-3-phosphate dehydrogenase ; Heat shock ; Heat shock proteins ; Human populations ; Identification ; Irrigation practices ; Leaves ; Middle East ; Muscle proteins ; Normalizing ; Panicum virgatum ; Pennisetum glaucum ; Phoeniceae - genetics ; Phoenix dactylifera ; Phosphates ; Physical Sciences ; Plant Leaves - genetics ; Plant Roots - genetics ; Population (statistical) ; Proline ; Proline transporter ; Rain ; Rainfall ; Real time ; Real-Time Polymerase Chain Reaction - methods ; Reference Standards ; Research and analysis methods ; Ribonucleic acid ; Ribosomal RNA ; RNA ; Roots ; rRNA 18S ; rRNA 25S ; Salinity ; Salinity effects ; Salinity tolerance ; Salts ; Seeds ; Semi arid areas ; Semiarid lands ; Semiarid zones ; Sodium Chloride ; Stability analysis ; Stress ; Stress analysis ; Stress, Physiological - genetics ; Superoxide dismutase ; Transporter ; Tubulin ; Ubiquitin ; Zinc</subject><ispartof>PloS one, 2016-11, Vol.11 (11), p.e0166216-e0166216</ispartof><rights>COPYRIGHT 2016 Public Library of Science</rights><rights>2016 V. Patankar et al. This is an open access article distributed under the terms of the Creative Commons Attribution License: http://creativecommons.org/licenses/by/4.0/ (the “License”), which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Notwithstanding the ProQuest Terms and Conditions, you may use this content in accordance with the terms of the License.</rights><rights>2016 V. Patankar et al 2016 V. Patankar et al</rights><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c626t-9ff63a4505899701adbaab76d362b098957e61b56c82ca1e43cd48cf661eedf93</citedby><cites>FETCH-LOGICAL-c626t-9ff63a4505899701adbaab76d362b098957e61b56c82ca1e43cd48cf661eedf93</cites><orcidid>0000-0002-5171-3014</orcidid></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktopdf>$$Uhttps://www.ncbi.nlm.nih.gov/pmc/articles/PMC5100987/pdf/$$EPDF$$P50$$Gpubmedcentral$$Hfree_for_read</linktopdf><linktohtml>$$Uhttps://www.ncbi.nlm.nih.gov/pmc/articles/PMC5100987/$$EHTML$$P50$$Gpubmedcentral$$Hfree_for_read</linktohtml><link.rule.ids>230,315,728,781,785,865,886,2103,2929,23868,27926,27927,53793,53795</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/27824922$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><contributor>Amancio, Sara</contributor><creatorcontrib>V Patankar, Himanshu</creatorcontrib><creatorcontrib>M Assaha, Dekoum V</creatorcontrib><creatorcontrib>Al-Yahyai, Rashid</creatorcontrib><creatorcontrib>Sunkar, Ramanjulu</creatorcontrib><creatorcontrib>Yaish, Mahmoud W</creatorcontrib><title>Identification of Reference Genes for Quantitative Real-Time PCR in Date Palm (Phoenix dactylifera L.) Subjected to Drought and Salinity</title><title>PloS one</title><addtitle>PLoS One</addtitle><description>Date palm is an important crop plant in the arid and semi-arid regions supporting human population in the Middle East and North Africa. These areas have been largely affected by drought and salinity due to insufficient rainfall and improper irrigation practices. Date palm is a relatively salt- and drought-tolerant plant and more recently efforts have been directed to identifying genes and pathways that confer stress tolerance in this species. Quantitative real-time PCR (qPCR) is a promising technique for the analysis of stress-induced differential gene expression, which involves the use of stable reference genes for normalizing gene expression. In an attempt to find the best reference genes for date palm's drought and salinity research, we evaluated the stability of 12 most commonly used reference genes using the geNorm, NormFinder, BestKeeper statistical algorithms and the comparative ΔCT method. The comprehensive results revealed that HEAT SHOCK PROTEIN (HSP), UBIQUITIN (UBQ) and YTH domain-containing family protein (YT521) were stable in drought-stressed leaves whereas GLYCERALDEHYDE-3-PHOSPHATE DEHYDROGENASE (GAPDH), ACTIN and TUBULIN were stable in drought-stressed roots. On the other hand, SMALL SUBUNIT RIBOSOMAL RNA (25S), YT521 and 18S ribosomal RNA (18S); and UBQ, ACTIN and ELONGATION FACTOR 1-ALPHA (eEF1a) were stable in leaves and roots, respectively, under salt stress. The stability of these reference genes was verified by using the abiotic stress-responsive CYTOSOLIC Cu/Zn SUPEROXIDE DISMUTASE (Cyt-Cu/Zn SOD), an ABA RECEPTOR, and a PROLINE TRANSPORTER 2 (PRO) genes. 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Academic</collection><collection>PubMed Central (Full Participant titles)</collection><collection>DOAJ Directory of Open Access Journals</collection><jtitle>PloS one</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>V Patankar, Himanshu</au><au>M Assaha, Dekoum V</au><au>Al-Yahyai, Rashid</au><au>Sunkar, Ramanjulu</au><au>Yaish, Mahmoud W</au><au>Amancio, Sara</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Identification of Reference Genes for Quantitative Real-Time PCR in Date Palm (Phoenix dactylifera L.) Subjected to Drought and Salinity</atitle><jtitle>PloS one</jtitle><addtitle>PLoS One</addtitle><date>2016-11-08</date><risdate>2016</risdate><volume>11</volume><issue>11</issue><spage>e0166216</spage><epage>e0166216</epage><pages>e0166216-e0166216</pages><issn>1932-6203</issn><eissn>1932-6203</eissn><abstract>Date palm is an important crop plant in the arid and semi-arid regions supporting human population in the Middle East and North Africa. These areas have been largely affected by drought and salinity due to insufficient rainfall and improper irrigation practices. Date palm is a relatively salt- and drought-tolerant plant and more recently efforts have been directed to identifying genes and pathways that confer stress tolerance in this species. Quantitative real-time PCR (qPCR) is a promising technique for the analysis of stress-induced differential gene expression, which involves the use of stable reference genes for normalizing gene expression. In an attempt to find the best reference genes for date palm's drought and salinity research, we evaluated the stability of 12 most commonly used reference genes using the geNorm, NormFinder, BestKeeper statistical algorithms and the comparative ΔCT method. The comprehensive results revealed that HEAT SHOCK PROTEIN (HSP), UBIQUITIN (UBQ) and YTH domain-containing family protein (YT521) were stable in drought-stressed leaves whereas GLYCERALDEHYDE-3-PHOSPHATE DEHYDROGENASE (GAPDH), ACTIN and TUBULIN were stable in drought-stressed roots. On the other hand, SMALL SUBUNIT RIBOSOMAL RNA (25S), YT521 and 18S ribosomal RNA (18S); and UBQ, ACTIN and ELONGATION FACTOR 1-ALPHA (eEF1a) were stable in leaves and roots, respectively, under salt stress. The stability of these reference genes was verified by using the abiotic stress-responsive CYTOSOLIC Cu/Zn SUPEROXIDE DISMUTASE (Cyt-Cu/Zn SOD), an ABA RECEPTOR, and a PROLINE TRANSPORTER 2 (PRO) genes. A combination of top 2 or 3 stable reference genes were found to be suitable for normalization of the target gene expression and will facilitate gene expression analysis studies aimed at identifying functional genes associated with drought and salinity tolerance in date palm.</abstract><cop>United States</cop><pub>Public Library of Science</pub><pmid>27824922</pmid><doi>10.1371/journal.pone.0166216</doi><orcidid>https://orcid.org/0000-0002-5171-3014</orcidid><oa>free_for_read</oa></addata></record> |
| fulltext | fulltext |
| identifier | ISSN: 1932-6203 |
| ispartof | PloS one, 2016-11, Vol.11 (11), p.e0166216-e0166216 |
| issn | 1932-6203 1932-6203 |
| language | eng |
| recordid | cdi_plos_journals_1837231344 |
| source | MEDLINE; DOAJ Directory of Open Access Journals; Public Library of Science (PLoS) Journals Open Access; EZB-FREE-00999 freely available EZB journals; PubMed Central; Free Full-Text Journals in Chemistry |
| subjects | Abiotic stress Actin Africa, Northern Algorithms Analysis Arid regions Arid zones Biology and Life Sciences Copper Drought Droughts Ecology and Environmental Sciences Elongation Gene expression Gene Expression Profiling - methods Gene Expression Regulation, Plant - genetics Genes Genes, Plant - genetics Genomes Glyceraldehyde-3-phosphate dehydrogenase Heat shock Heat shock proteins Human populations Identification Irrigation practices Leaves Middle East Muscle proteins Normalizing Panicum virgatum Pennisetum glaucum Phoeniceae - genetics Phoenix dactylifera Phosphates Physical Sciences Plant Leaves - genetics Plant Roots - genetics Population (statistical) Proline Proline transporter Rain Rainfall Real time Real-Time Polymerase Chain Reaction - methods Reference Standards Research and analysis methods Ribonucleic acid Ribosomal RNA RNA Roots rRNA 18S rRNA 25S Salinity Salinity effects Salinity tolerance Salts Seeds Semi arid areas Semiarid lands Semiarid zones Sodium Chloride Stability analysis Stress Stress analysis Stress, Physiological - genetics Superoxide dismutase Transporter Tubulin Ubiquitin Zinc |
| title | Identification of Reference Genes for Quantitative Real-Time PCR in Date Palm (Phoenix dactylifera L.) Subjected to Drought and Salinity |
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