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金华猪和大约克猪泌乳期乳腺组织的microRNA和转录组测序分析.docx

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金华猪和大约克猪泌乳期乳腺组织的microRNA和转录组测序分析 Introduction Pigsareimportantlivestockthatprovidemeatandby-productssuchaslardandleather.Theproductivityofpigsdependsontheirhealth,nutrition,andreproductiveperformance.Oneoftheimportantparametersformeasuringreproductiveperformanceinpigsisthemilkproductionduringlactation,whichiscriticalforthesurvivalandgrowthofpiglets.Milkproductionisregulatedbycomplexmolecularmechanismsinvolvingvariousbiomolecules,includingmicroRNAsandgenes.Inthisstudy,weanalyzedthemicroRNAsandtranscriptomesofthemammaryglandsoftwopigbreeds,JinhuaandDurocYork,duringlactationtogaininsightsintothemolecularmechanismsinvolvedinmilkproduction. MaterialsandMethods Animals Ten2-year-oldsowsfromeachbreedwereselectedforthisstudy.Theywerehousedinthesamefacility,fedthesamediet,andtheirhealthwasmonitored. MammaryglandsamplingandRNAextraction Mammaryglandtissuewascollectedfromeachsowat1,7,and21daysafterfarrowing.Thesampleswereimmediatelysnap-frozeninliquidnitrogenandstoredat-80°Cuntilanalysis.TotalRNAwasextractedusingTRIzolreagentaccordingtothemanufacturer'sinstructions. MicroRNAsequencing MicroRNAlibrarieswerepreparedusingtheNEBNextMultiplexSmallRNALibraryPrepSetforIlluminasequencing(NewEnglandBiolabs).SequencingwascarriedoutontheIlluminaHiSeqXTenplatformwithareadlengthof50bp. Transcriptomesequencing RNAsequencinglibrarieswerepreparedusingtheNEBNextUltraRNALibraryPrepKitforIlluminasequencing(NewEnglandBiolabs).SequencingwascarriedoutontheIlluminaHiSeqXTenplatformwithareadlengthof150bp. Dataanalysis TherawsequencingdatawereprocessedusingFastQCtogeneratequalityreports.Theadaptersandlow-qualityreadswereremovedusingTrimmomatic.Thecleanreadsweremappedtothepiggenome(Susscrofa11.1)usingBowtie2.TheexpressionlevelsofmicroRNAsandgeneswerequantifiedusingthefeatureCountsprogram.DifferentialexpressionanalysiswasperformedusingtheDESeq2packageinR.Geneontology(GO)andKyotoEncyclopediaofGenesandGenomes(KEGG)pathwayanalyseswereperformedusingtheDAVIDandKOBASdatabases,respectively. Results MicroRNAseq