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Highlight
red
red

<P5 primer/capture site>

Highlight
yellow
yellow

<IndexRead2>

Highlight
green
green

<Read1 primer site>

<template - gDNA, RNA, amplicon, whatever>
Highlight
cyan
cyan

<Read2 primer site>

Highlight
blue
blue

<IndexRead1>

Highlight
purple
purple

<P7 primer/capture site>

Note: Highlighting provided prior to 6/2012 was incorrect. It showed the Index or Barcode Read 2 sequence as GATCT when in fact the second index sequence is downstream of that site. We apologize for the error. See the section entitled, "Dual-index adaptor design..." below for corrected documentationIf you'd like a different description, this one from the Tufts core facility is quite good.

Single index adaptor design on a standard Illumina HiSeq or MiSeq run

...

Code Block
Oligonucleotide sequences for TruSeqTM RNA and DNA Sample Prep Kits1

TruSeq Universal Adapter
     5’ AATGATACGGCGACCACCGAGATCTACACTCTTTCCCTACACGACGCTCTTCCGATCT

TruSeq Adapters
barcode:                            ATCACG
TruSeq Adapter, Index 1
5’ GATCGGAAGAGCACACGTCTGAACTCCAGTCACATCACGATCTCGTATGCCGTCTTCTGCTTG
TruSeq Adapter, Index 2
5’ GATCGGAAGAGCACACGTCTGAACTCCAGTCACCGATGTATCTCGTATGCCGTCTTCTGCTTG
TruSeq Adapter, Index 3
5’ GATCGGAAGAGCACACGTCTGAACTCCAGTCACTTAGGCATCTCGTATGCCGTCTTCTGCTTG
TruSeq Adapter, Index 4
5’ GATCGGAAGAGCACACGTCTGAACTCCAGTCACTGACCAATCTCGTATGCCGTCTTCTGCTTG
TruSeq Adapter, Index 5
5’ GATCGGAAGAGCACACGTCTGAACTCCAGTCACACAGTGATCTCGTATGCCGTCTTCTGCTTG
TruSeq Adapter, Index 6
5’ GATCGGAAGAGCACACGTCTGAACTCCAGTCACGCCAATATCTCGTATGCCGTCTTCTGCTTG
TruSeq Adapter, Index 7
5’ GATCGGAAGAGCACACGTCTGAACTCCAGTCACCAGATCATCTCGTATGCCGTCTTCTGCTTG
TruSeq Adapter, Index 8
5’ GATCGGAAGAGCACACGTCTGAACTCCAGTCACACTTGAATCTCGTATGCCGTCTTCTGCTTG
TruSeq Adapter, Index 9
5’ GATCGGAAGAGCACACGTCTGAACTCCAGTCACGATCAGATCTCGTATGCCGTCTTCTGCTTG
TruSeq Adapter, Index 10
5’ GATCGGAAGAGCACACGTCTGAACTCCAGTCACTAGCTTATCTCGTATGCCGTCTTCTGCTTG
TruSeq Adapter, Index 11
5’ GATCGGAAGAGCACACGTCTGAACTCCAGTCACGGCTACATCTCGTATGCCGTCTTCTGCTTG
TruSeq Adapter, Index 12
5’ GATCGGAAGAGCACACGTCTGAACTCCAGTCACCTTGTAATCTCGTATGCCGTCTTCTGCTTG
5' GATCGGAAGAGCACACGTCTGAACTCCAGTCAC Mux index read seq primer
  AGATCGGAAGAGCACACGTCTG - my 3' adaptor
my RT primer:
5' TCAGACGTGTGCTCTTCCGATCT 3'
  TCTAGCCTTCTCGTGTGCAGACTTGAGGTCAGTG Mux read 2 seq primer (reversed)
                       CTTGAGGTCAGTGGAACATTAGAGCATACGGCAGAAGACGAAC Index 12 PCR primer (reversed)
So order RC of TruSeq Adaptor Indexes as PCR primers

Oligonucleotide sequences for TruSeq Small RNA Sample Prep Kits
*****************************
Example (all Illumina sequences unless noted):
RNA PCR Primer (RP1),  part # 15005505
5’ AATGATACGGCGACCACCGAGATCTACACGTTCAGAGTTCTACAGTCCGA
   AATGATACGGCGACCACCGA      CAGGTTCAGAGTTCTACAGTCCGA - this is the NEB SR Primer R1 - pads added by SPHS
                 5' RNA adaptor GUUCAGAGUUCUACAGUCCGACGAUC  (NEB kit calls this the SR Adaptor 1)

RNA PCR Primer, Index 1 (RPI1) code ATCACG in DNA, RC (CGTGAT)
5' CAAGCAGAAGACGGCATACGAGATCGTGATGTGACTGGAGTTCCTTGGCACCCGAGAATTCCA 3' - RNA pcr primer
   CAAGCAGAAGACGGCATACGAGATCGTGATGTGACTGGAGTTCAGACGTGTGCTCTTCCGATC (RC of DNA Index 1 for comparison)
                        3' RNA adaptor   3'  CCTTGGCACCCGAGAATTCCA - 5'
*****************************
Differences with NEB Small RNA kit (RP1/SR Primer R1/SR Adaptor 1/5' RNA adaptor are all the same):
5’ GATCGGAAGAGCACACGTCTGAACTCCAGTCACCTTGTAATCTCGTATGCCGTCTTCTGCTTG  (TruSeq Adaptor Ind 12)
 NEB 3' adaptor (3' SR Adaptor 1)       5'  ATCGTATGCCGTCTTCTGCTTG 3'
                                       3'    AGCATACGGCAGAAGACGAAC 5' (reverse of NEB SR Primer F1)
3' CTAGCCTTCTCGTGTGCAGACTTGAGGTCAGTGCCGATGTAGAGCATACGGCAGAAGACGAAC 5' (rev. of Ind. 12 adaptor)
CAAGCAGAAGACGGCATACGAGATTACAAGGTGACTGGAGTTC (TruSeq Adaptor Ind 12 PCR primer)
5' CAAGCAGAAGACGGCATACGA 3' (NEB SR Primer F1)
5' CAAGCAGAAGACGGCATACGAGCTCTTCCGATCT   (Illumina genomic PCR primer)
3' CAAGCAGAAGACGGCATACGAT 5' - NEB 3' SR Adaptor 1 (RC)
5' CAAGCAGAAGACGGCATACGA 3' - NEB RT Primer 1
For reference:
>TruSeqAdapterIndex11_RC
CAAGCAGAAGACGGCATACGAGATGTAGCCGTGACTGGAGTTCAGACGTGTGCTCTTCCGATC
>TruSeqAdapterIndex12_RC
CAAGCAGAAGACGGCATACGAGATTACAAGGTGACTGGAGTTCAGACGTGTGCTCTTCCGATC
*****************************

RNA 5’ Adapter (RA5),  part # 15013205
5’ GUUCAGAGUUCUACAGUCCGACGAUC

RNA 3’ Adapter (RA3),  part # 15013207
5’ TGGAATTCTCGGGTGCCAAGG
3' ACCTTAAGAGCCCACGGTTCCG 5' (RT primer, reversed)

RNA RT Primer (RTP),  part # 15013981
5’ GCCTTGGCACCCGAGAATTCCA

Multiplexing Index Read Sequencing Primer
5' GATCGGAAGAGCACACGTCTGAACTCCAGTCAC
Multiplexing Read 2 Sequencing Primer
5' GTGACTGGAGTTCAGACGTGTGCTCTTCCGATCT
RNA PCR Primer (RP1),  part # 15005505
5’ AATGATACGGCGACCACCGAGATCTACACGTTCAGAGTTCTACAGTCCGA
RNA PCR Primer, Index 1 (RPI1)
CAAGCAGAAGACGGCATACGAGATCGTGATGTGACTGGAGTTCCTTGGCACCCGAGAATTCCA
RNA PCR Primer, Index 2 (RPI2)
CAAGCAGAAGACGGCATACGAGATACATCGGTGACTGGAGTTCCTTGGCACCCGAGAATTCCA
RNA PCR Primer, Index 3 (RPI3)
CAAGCAGAAGACGGCATACGAGATGCCTAAGTGACTGGAGTTCCTTGGCACCCGAGAATTCCA
RNA PCR Primer, Index 4 (RPI4)
CAAGCAGAAGACGGCATACGAGATTGGTCAGTGACTGGAGTTCCTTGGCACCCGAGAATTCCA
RNA PCR Primer, Index 5 (RPI5)
CAAGCAGAAGACGGCATACGAGATCACTGTGTGACTGGAGTTCCTTGGCACCCGAGAATTCCA
RNA PCR Primer, Index 6 (RPI6)
CAAGCAGAAGACGGCATACGAGATATTGGCGTGACTGGAGTTCCTTGGCACCCGAGAATTCCA
RNA PCR Primer, Index 7 (RPI7)
CAAGCAGAAGACGGCATACGAGATGATCTGGTGACTGGAGTTCCTTGGCACCCGAGAATTCCA
RNA PCR Primer, Index 8 (RPI8)
CAAGCAGAAGACGGCATACGAGATTCAAGTGTGACTGGAGTTCCTTGGCACCCGAGAATTCCA
RNA PCR Primer, Index 9 (RPI9)
CAAGCAGAAGACGGCATACGAGATCTGATCGTGACTGGAGTTCCTTGGCACCCGAGAATTCCA
RNA PCR Primer, Index 10 (RPI10)
CAAGCAGAAGACGGCATACGAGATAAGCTAGTGACTGGAGTTCCTTGGCACCCGAGAATTCCA
RNA PCR Primer, Index 11 (RPI11)
CAAGCAGAAGACGGCATACGAGATGTAGCCGTGACTGGAGTTCCTTGGCACCCGAGAATTCCA
RNA PCR Primer, Index 12 (RPI12)
CAAGCAGAAGACGGCATACGAGATTACAAGGTGACTGGAGTTCCTTGGCACCCGAGAATTCCA
RNA PCR Primer, Index 13 (RPI13)
CAAGCAGAAGACGGCATACGAGATTTGACTGTGACTGGAGTTCCTTGGCACCCGAGAATTCCA
RNA PCR Primer, Index 14 (RPI14)
CAAGCAGAAGACGGCATACGAGATGGAACTGTGACTGGAGTTCCTTGGCACCCGAGAATTCCA
RNA PCR Primer, Index 15 (RPI15)
CAAGCAGAAGACGGCATACGAGATTGACATGTGACTGGAGTTCCTTGGCACCCGAGAATTCCA
RNA PCR Primer, Index 16 (RPI16)
CAAGCAGAAGACGGCATACGAGATGGACGGGTGACTGGAGTTCCTTGGCACCCGAGAATTCCA
RNA PCR Primer, Index 17 (RPI17)
CAAGCAGAAGACGGCATACGAGATCTCTACGTGACTGGAGTTCCTTGGCACCCGAGAATTCCA
RNA PCR Primer, Index 18 (RPI18)
CAAGCAGAAGACGGCATACGAGATGCGGACGTGACTGGAGTTCCTTGGCACCCGAGAATTCCA
RNA PCR Primer, Index 19 (RPI19)
CAAGCAGAAGACGGCATACGAGATTTTCACGTGACTGGAGTTCCTTGGCACCCGAGAATTCCA
RNA PCR Primer, Index 20 (RPI20)
CAAGCAGAAGACGGCATACGAGATGGCCACGTGACTGGAGTTCCTTGGCACCCGAGAATTCCA
RNA PCR Primer, Index 21 (RPI21)
CAAGCAGAAGACGGCATACGAGATCGAAACGTGACTGGAGTTCCTTGGCACCCGAGAATTCCA
RNA PCR Primer, Index 22 (RPI22)
CAAGCAGAAGACGGCATACGAGATCGTACGGTGACTGGAGTTCCTTGGCACCCGAGAATTCCA
RNA PCR Primer, Index 23 (RPI23)
CAAGCAGAAGACGGCATACGAGATCCACTCGTGACTGGAGTTCCTTGGCACCCGAGAATTCCA
RNA PCR Primer, Index 24 (RPI24)
CAAGCAGAAGACGGCATACGAGATGCTACCGTGACTGGAGTTCCTTGGCACCCGAGAATTCCA
RNA PCR Primer, Index 25 (RPI25)
CAAGCAGAAGACGGCATACGAGATATCAGTGTGACTGGAGTTCCTTGGCACCCGAGAATTCCA
RNA PCR Primer, Index 26 (RPI26)
CAAGCAGAAGACGGCATACGAGATGCTCATGTGACTGGAGTTCCTTGGCACCCGAGAATTCCA
RNA PCR Primer, Index 27 (RPI27)
CAAGCAGAAGACGGCATACGAGATAGGAATGTGACTGGAGTTCCTTGGCACCCGAGAATTCCA
RNA PCR Primer, Index 28 (RPI28)
CAAGCAGAAGACGGCATACGAGATCTTTTGGTGACTGGAGTTCCTTGGCACCCGAGAATTCCA
RNA PCR Primer, Index 29 (RPI29)
CAAGCAGAAGACGGCATACGAGATTAGTTGGTGACTGGAGTTCCTTGGCACCCGAGAATTCCA
RNA PCR Primer, Index 30 (RPI30)
CAAGCAGAAGACGGCATACGAGATCCGGTGGTGACTGGAGTTCCTTGGCACCCGAGAATTCCA
RNA PCR Primer, Index 31 (RPI31)
CAAGCAGAAGACGGCATACGAGATATCGTGGTGACTGGAGTTCCTTGGCACCCGAGAATTCCA
RNA PCR Primer, Index 32 (RPI32)
CAAGCAGAAGACGGCATACGAGATTGAGTGGTGACTGGAGTTCCTTGGCACCCGAGAATTCCA
RNA PCR Primer, Index 33 (RPI33)
CAAGCAGAAGACGGCATACGAGATCGCCTGGTGACTGGAGTTCCTTGGCACCCGAGAATTCCA
RNA PCR Primer, Index 34 (RPI34)
CAAGCAGAAGACGGCATACGAGATGCCATGGTGACTGGAGTTCCTTGGCACCCGAGAATTCCA
RNA PCR Primer, Index 35 (RPI35)
CAAGCAGAAGACGGCATACGAGATAAAATGGTGACTGGAGTTCCTTGGCACCCGAGAATTCCA
RNA PCR Primer, Index 36 (RPI36)
CAAGCAGAAGACGGCATACGAGATTGTTGGGTGACTGGAGTTCCTTGGCACCCGAGAATTCCA
RNA PCR Primer, Index 37 (RPI37)
CAAGCAGAAGACGGCATACGAGATATTCCGGTGACTGGAGTTCCTTGGCACCCGAGAATTCCA
RNA PCR Primer, Index 38 (RPI38)
CAAGCAGAAGACGGCATACGAGATAGCTAGGTGACTGGAGTTCCTTGGCACCCGAGAATTCCA
RNA PCR Primer, Index 39 (RPI39)
CAAGCAGAAGACGGCATACGAGATGTATAGGTGACTGGAGTTCCTTGGCACCCGAGAATTCCA
RNA PCR Primer, Index 40 (RPI40)
CAAGCAGAAGACGGCATACGAGATTCTGAGGTGACTGGAGTTCCTTGGCACCCGAGAATTCCA
RNA PCR Primer, Index 41 (RPI41)
CAAGCAGAAGACGGCATACGAGATGTCGTCGTGACTGGAGTTCCTTGGCACCCGAGAATTCCA
RNA PCR Primer, Index 42 (RPI42)
CAAGCAGAAGACGGCATACGAGATCGATTAGTGACTGGAGTTCCTTGGCACCCGAGAATTCCA
RNA PCR Primer, Index 43 (RPI43)
CAAGCAGAAGACGGCATACGAGATGCTGTAGTGACTGGAGTTCCTTGGCACCCGAGAATTCCA
RNA PCR Primer, Index 44 (RPI44)
CAAGCAGAAGACGGCATACGAGATATTATAGTGACTGGAGTTCCTTGGCACCCGAGAATTCCA
RNA PCR Primer, Index 45 (RPI45)
CAAGCAGAAGACGGCATACGAGATGAATGAGTGACTGGAGTTCCTTGGCACCCGAGAATTCCA
RNA PCR Primer, Index 46 (RPI46)
CAAGCAGAAGACGGCATACGAGATTCGGGAGTGACTGGAGTTCCTTGGCACCCGAGAATTCCA
RNA PCR Primer, Index 47 (RPI47)
CAAGCAGAAGACGGCATACGAGATCTTCGAGTGACTGGAGTTCCTTGGCACCCGAGAATTCCA
RNA PCR Primer, Index 48 (RPI48)
CAAGCAGAAGACGGCATACGAGATTGCCGAGTGACTGGAGTTCCTTGGCACCCGAGAATTCCA


Oligonucleotide sequences for Genomic DNA
 PCR Primer, Index 48 (RPI48)
CAAGCAGAAGACGGCATACGAGATTGCCGAGTGACTGGAGTTCCTTGGCACCCGAGAATTCCA


Oligonucleotide sequences for Genomic DNA

Adapters
5' P-GATCGGAAGAGCTCGTATGCCGTCTTCTGCTTG
5' ACACTCTTTCCCTACACGACGCTCTTCCGATCT
PCR Primers
5' AATGATACGGCGACCACCGAGATCTACACTCTTTCCCTACACGACGCTCTTCCGATCT
5' CAAGCAGAAGACGGCATACGAGCTCTTCCGATCT

Genomic DNA Sequencing Primer
5' ACACTCTTTCCCTACACGACGCTCTTCCGATCT


Paired End DNA oligonucleotide sequences

PE Adapters
5' P-GATCGGAAGAGCTCGTATGCCGTCTTCTGCTTGGATCGGAAGAGCGGTTCAGCAGGAATGCCGAG
5' ACACTCTTTCCCTACACGACGCTCTTCCGATCT
PE PCR Primer Primers1.0
5' AATGATACGGCGACCACCGAGATCTACACTCTTTCCCTACACGACGCTCTTCCGATCT
PE PCR Primer 2.0
5' CAAGCAGAAGACGGCATACGAGATCGGTCTCGGCATTCCTGCTGAACCGCTCTTCCGATCT

PE Read 1 Sequencing Primer
5' CAAGCAGAAGACGGCATACGAGCTCTTCCGATCTACACTCTTTCCCTACACGACGCTCTTCCGATCT
PE GenomicRead DNA2 Sequencing Primer
5' ACACTCTTTCCCTACACGACGCTCTTCCGATCT


Paired End DNA oligonucleotide sequences

PE CGGTCTCGGCATTCCTGCTGAACCGCTCTTCCGATCT


Oligonucleotide sequences for the Multiplexing Sample Prep Oligo Only Kit

Multiplexing Adapters
5' P-GATCGGAAGAGCGGTTCAGCAGGAATGCCGAGGATCGGAAGAGCACACGTCT
5' ACACTCTTTCCCTACACGACGCTCTTCCGATCT
PEMultiplexing PCR Primer 1.0
5' AATGATACGGCGACCACCGAGATCTACACTCTTTCCCTACACGACGCTCTTCCGATCT
PEMultiplexing PCR Primer 2.0
5' CAAGCAGAAGACGGCATACGAGATCGGTCTCGGCATTCCTGCTGAACCGCTCTTCCGATCT

PE
Multiplexing Read 1 Sequencing Primer
5' ACACTCTTTCCCTACACGACGCTCTTCCGATCT
PE Read 2 Sequencing Primer
5' CGGTCTCGGCATTCCTGCTGAACCGCTCTTCCGATCT


Oligonucleotide sequences for the ACACTCTTTCCCTACACGACGCTCTTCCGATCT
Multiplexing SampleIndex PrepRead OligoSequencing OnlyPrimer
Kit5' GATCGGAAGAGCACACGTCTGAACTCCAGTCAC
Multiplexing Read Adapters2 5' P-GATCGGAAGAGCACACGTCTSequencing Primer
5' ACACTCTTTCCCTACACGACGCTCTTCCGATCTGTGACTGGAGTTCAGACGTGTGCTCTTCCGATCT
Multiplexing
PCR Primer, Index 1.0
5' AATGATACGGCGACCACCGAGATCTACACTCTTTCCCTACACGACGCTCTTCCGATCT
Multiplexing
CAAGCAGAAGACGGCATACGAGATCGTGATGTGACTGGAGTTC
PCR Primer, Index 2.0
CAAGCAGAAGACGGCATACGAGATACATCGGTGACTGGAGTTC
MultiplexingPCR ReadPrimer, 1Index Sequencing3
PrimerCAAGCAGAAGACGGCATACGAGATGCCTAAGTGACTGGAGTTC
5' ACACTCTTTCCCTACACGACGCTCTTCCGATCT
MultiplexingPCR Primer, Index 4
ReadCAAGCAGAAGACGGCATACGAGATTGGTCAGTGACTGGAGTTC
SequencingPCR Primer, Index 5'
GATCGGAAGAGCACACGTCTGAACTCCAGTCAC
Multiplexing Read 2 SequencingCAAGCAGAAGACGGCATACGAGATCACTGTGTGACTGGAGTTC
PCR Primer, 5'Index GTGACTGGAGTTCAGACGTGTGCTCTTCCGATCT6
CAAGCAGAAGACGGCATACGAGATATTGGCGTGACTGGAGTTC
PCR Primer, Index 17
CAAGCAGAAGACGGCATACGAGATCGTGATGTGACTGGAGTTCCAAGCAGAAGACGGCATACGAGATGATCTGGTGACTGGAGTTC
PCR Primer, Index 28
CAAGCAGAAGACGGCATACGAGATACATCGGTGACTGGAGTTCCAAGCAGAAGACGGCATACGAGATTCAAGTGTGACTGGAGTTC
PCR Primer, Index 39
CAAGCAGAAGACGGCATACGAGATGCCTAAGTGACTGGAGTTCCAAGCAGAAGACGGCATACGAGATCTGATCGTGACTGGAGTTC
PCR Primer, Index 410
CAAGCAGAAGACGGCATACGAGATTGGTCAGTGACTGGAGTTCCAAGCAGAAGACGGCATACGAGATAAGCTAGTGACTGGAGTTC
PCR Primer, Index 511
CAAGCAGAAGACGGCATACGAGATCACTGTGTGACTGGAGTTCCAAGCAGAAGACGGCATACGAGATGTAGCCGTGACTGGAGTTC
PCR Primer, Index 6
CAAGCAGAAGACGGCATACGAGATATTGGCGTGACTGGAGTTC
PCR Primer, Index 7
CAAGCAGAAGACGGCATACGAGATGATCTGGTGACTGGAGTTC
PCR Primer, Index 8
CAAGCAGAAGACGGCATACGAGATTCAAGTGTGACTGGAGTTC
PCR Primer, Index 9
CAAGCAGAAGACGGCATACGAGATCTGATCGTGACTGGAGTTC
PCR Primer, Index 10
CAAGCAGAAGACGGCATACGAGATAAGCTAGTGACTGGAGTTC
PCR Primer, Index 11
CAAGCAGAAGACGGCATACGAGATGTAGCCGTGACTGGAGTTC
PCR Primer, Index 12
CAAGCAGAAGACGGCATACGAGATTACAAGGTGACTGGAGTTC 12
CAAGCAGAAGACGGCATACGAGATTACAAGGTGACTGGAGTTC

Oligonucleotide sequences © 2007-2012 Illumina, Inc. All rights reserved. 
Derivative works created by Illumina customers are authorized for use with Illumina 
instruments and products only. All other uses are strictly prohibited.