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转录组测序建库流程
英文回答：
Transcriptomesequencing,alsoknownasRNA-Seq,isa
powerfultoolusedtostudytheexpressionandregulation
ofgenesinanorganism.Itinvolvestheconstructionofa
cDNAlibraryfromRNAsamples,whichisthensequenced
usinghigh-throughputsequencingtechnologies.Theprocess
oftranscriptomesequencinginvolvesseveralsteps,
includingRNAextraction,librarypreparation,sequencing,
anddataanalysis.
Firstly,RNAextractionisperformedtoisolatetheRNA
moleculesfromthebiologicalsampleofinterest.Thiscan
bedoneusingvariousmethods,suchasphenol-chloroform
extractionorcolumn-basedpurificationkits.Thequality
andintegrityoftheRNAarecrucialforthesuccessofthe
subsequentsteps.
Next,theextractedRNAisconvertedintocDNAthrough
aprocesscalledreversetranscription.Thisinvolvesthe
useofreversetranscriptaseenzymestosynthesize
complementaryDNA(cDNA)fromtheRNAtemplate.ThecDNAis
thenfragmentedintosmallerpiecestofacilitate
sequencing.
Afterfragmentation,adaptersareaddedtothecDNA
fragments.Theseadapterscontainsequencesthatare
recognizedbythesequencingplatformandenablethe
attachmentofthefragmentstothesequencingflowcell.
Thisstepisessentialforthesubsequentsequencing
process.
Oncethelibrarypreparationiscomplete,thecDNA
fragmentsarereadyforsequencing.High-throughput
sequencingplatforms,suchasIlluminaorIonTorrent,are
commonlyusedfortranscriptomesequencing.Theseplatforms
generatemillionsofshortreadsthatrepresentthecDNA
fragments.
Thegeneratedsequencingdatathenundergoesaseries
ofbioinformaticsanalyses.Thisincludesqualitycontrol,
readalignmenttoareferencegenomeortranscriptome,and
quantificationofgeneexpressionlevels.Varioussoftware
toolsandpipelinesareavailablefortheseanalyses,such
asFastQC,STAR,andDESeq2.
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