{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"institution":[{"name":"Authorea, Inc."}],"indexed":{"date-parts":[[2025,6,3]],"date-time":"2025-06-03T16:38:49Z","timestamp":1748968729116,"version":"3.41.0"},"posted":{"date-parts":[[2022,7,25]]},"group-title":"Preprints","reference-count":0,"publisher":"Wiley","content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"accepted":{"date-parts":[[2022,7,25]]},"abstract":"<jats:p id=\"p1\">RNA sequencing (RNA-Seq) is a popular method for measuring gene\nexpression in non-model organisms, including wild populations. While\nRNA-Seq can measure gene expression variation among wild-caught\nindividuals and can yield important biological insights into organism\nfunction, sampling methods may also influence gene expression estimates.\nWe examined the influence of multiple technical variables on estimated\ngene expression in a non-model fish, the westslope cutthroat trout\n(Oncorhynchus clarkii lewisi), using two RNA-Seq library types: 3\u2019\nRNA-Seq and whole mRNA-Seq. We evaluated effects of dip netting versus\nelectrofishing, and of harvesting tissue immediately versus 5 minutes\nafter euthanasia on estimated gene expression in blood, gill, and\nmuscle. We detected 30% more genes with whole mRNA-Seq than with 3\u2019\nRNA-Seq and found that 58% of genes were significantly differently\nexpressed between 3\u2019 RNA-Seq and whole mRNA-Seq. Our findings indicate\nthat 3\u2019 RNA-Seq and whole mRNA-Seq are robust to the technical variables\nrelated to the field sampling approaches tested here with a lack of\ndifferential gene expression among sampling methods and tissue\ncollection time after euthanasia. However, we found that gene expression\nvaried based on which RNA-Seq library type was used on the same set of\nsamples. Our study suggests researchers could safely rely on different\nfish sampling strategies in the field and save money and analyze more\nindividuals using 3\u2019 RNA-Seq, but should use whole mRNA-Seq when working\nwith a species without good genomic resources, and when maximizing the\nnumber of genes identified and detecting alternative splicing are\nimportant.<\/jats:p>","DOI":"10.22541\/au.165875386.65530784\/v1","type":"posted-content","created":{"date-parts":[[2022,7,25]],"date-time":"2022-07-25T12:57:54Z","timestamp":1658753874000},"source":"Crossref","is-referenced-by-count":0,"title":["Influence of RNA-Seq library construction, sampling methods, and tissue harvesting time on gene expression estimation"],"prefix":"10.22541","author":[{"ORCID":"https:\/\/orcid.org\/0000-0003-2338-8602","authenticated-orcid":true,"given":"Ylenia","family":"Chiari","sequence":"first","affiliation":[{"name":"George Mason University"}]},{"given":"Leif","family":"Howard","sequence":"additional","affiliation":[{"name":"University of Montana Missoula Division of Biological Sciences"}]},{"given":"Nickolas","family":"Moreno","sequence":"additional","affiliation":[{"name":"George Mason University"}]},{"given":"Scott","family":"Relyea","sequence":"additional","affiliation":[{"name":"Montana Fish Wildlife and Parks"}]},{"given":"James","family":"Dunnigan","sequence":"additional","affiliation":[{"name":"Montana Fish Wildlife and Parks"}]},{"given":"Matthew","family":"Boyer","sequence":"additional","affiliation":[{"name":"University of Montana"}]},{"given":"Marty","family":"Kardos","sequence":"additional","affiliation":[{"name":"National Oceanic and Atmospheric Center"}]},{"given":"Scott","family":"Glaberman","sequence":"additional","affiliation":[{"name":"George Mason University"}]},{"given":"Gordon","family":"Luikart","sequence":"additional","affiliation":[{"name":"University of Montana Missoula Division of Biological Sciences"}]}],"member":"311","container-title":[],"original-title":[],"deposited":{"date-parts":[[2022,7,25]],"date-time":"2022-07-25T12:57:54Z","timestamp":1658753874000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.authorea.com\/users\/391305\/articles\/578403-influence-of-rna-seq-library-construction-sampling-methods-and-tissue-harvesting-time-on-gene-expression-estimation?commit=c6cc87bf50b97ecd80cbff98ce39980818adee54"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2022,7,25]]},"references-count":0,"URL":"https:\/\/doi.org\/10.22541\/au.165875386.65530784\/v1","relation":{},"subject":[],"published":{"date-parts":[[2022,7,25]]},"subtype":"preprint"}}