Description:


TheNEBNext® Ultra™DNALibraryPrepKitforIlluminacontainsenzymesandbuffersthatareidealtoconvertasmallamountofDNAinputintoindexedlibrariesfornext-generationsequencingontheIlluminaplatform(Illumina,Inc).TheworkflowofNEBNextUltraDNALibraryPrepKitforIlluminaisveryuser-friendlyandfastwithminimalhands-ontime.Eachofthesecomponentsmustpassrigorousqualitycontrolstandardsandarelotcontrolled,bothindividuallyandasasetofreagents.
UltraDNALibraryPreparationWorkflowforIllumina
NEBNextUltra™DNAprovideshighlibraryyieldsevenwithlow inputsanddifficultsamples
Alllibrarieswerepreparedwith5ngofinputDNAandrunonanAgilentBioanalyzer®.
A.E.coligenomicDNAlibraries.1:Ladder;2:LibrarypreparedusingNEBNextDNA LibraryPrepMasterMixSet(E6040);3:LibrarypreparedusingNEBNextUltraDNA LibraryPrepKit(E7370).With5nginputs,libraryyieldsaresignificantlyhigherusing theNEBNextUltra™DNAKit.
B.FFPEsample.HumanlungtumorgenomicDNAfroma66-yearoldmale,extractedfrom a10-yearoldFFPEblock(Biochain,Inc.).1:LibrarypreparedusingNEBNextUltraDNA LibraryPrepKit(E7370).2:InputDNA(highlydegradedFFPEgenomicDNA).Readdepthcorrelationshowsconsistenthighcoveragefor5ng-1µginputamounts
LibrarieswerepreparedwithvariousamountsofhumanIMR90gDNA(5,50,and1000ng)andPCRamplified.APCR-freelibrarywasalsopreparedwith1000nghumanIMR90gDNA.Sequencecoverageof10kbwindowsofthehumangenomewasanalyzedonGalaxy,andPearsoncorrelationcoefficient(R2values)werecalculatedusingMicrosoft®Excel®.Alllibrariesarehighlycorrelatedwitheachother,independentofDNAinputaswellasPCRamplification.HighcoveragewasachievedforallDNAinputamounts.
NEBNextUltraDNAKitsdisplaystrongperformancewithinputamountsaslowas5ng

InputDNA:IMR90genomicDNA(humanfibroblast).
PFMismatchRate:Therateofbasesmismatchingthereferenceforallbasesalignedtothereferencesequence.
PercentDuplication:Thepercentageofmappedsequencethatismarkedasduplicate.
PercentChimeras:Thepercentageofreadsthatmapoutsideofamaximuminsertsizeorthathavethetwoendsmappingtodifferentchromosomes.
ThisdataillustratesthestrongperformanceoftheNEBNextUltraDNALibraryPrepKitforIllumina,evenwith5ngofhumangenomicDNAinput.Mismatchrate,%duplicationand%chimerasarealllow,andsimilar,regardlessofinputamount.
FunctionalValidation
EachsetofreagentsisfunctionallyvalidatedtogetherthroughconstructionandsequencingofaDNAlibraryontheIlluminasequencingplatform.LotControl
Thelotsprovidedaremanagedseparatelyandqualifiedbyadditionalfunctionalvalidation.Individualreagentsundergostandardenzymeactivityandqualitycontrolassays,andalsomeetstringentcriteriaintheadditionalqualitycontrolslistedoneachindividualcomponentpage
ReagentsSupplied
Thefollowingreagentsaresuppliedwiththisproduct:
| Storeat(°C) | Concentration |
| NEBNextEndPrepEnzymeMix | -20 | |
| NEBNextEndRepairReactionBuffer | -20 | 10X |
| Blunt/TALigaseMasterMix | -20 | |
| NEBNext®LigationEnhancer | -20 | |
| NEBNext®Q5®HotStartHiFiPCRMasterMix | -20 | |