Illumina GA/Oligo info: Difference between revisions
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| TruS_R||5'- CAAGCAGAAGACGGCATACGAGAT -3'||Tm = 65.5 C | | TruS_R||5'- CAAGCAGAAGACGGCATACGAGAT -3'||Tm = 65.5 C | ||
|} | |} | ||
'''Read1:''' HP10<br> | |||
'''In dex read:'''HP12<br> | |||
'''Read2:''' HP11 |
Revision as of 20:45, 17 May 2014
Name in original paper | Nick-name | Sequence (5\'->3\') | Sequence Modification | Reference | |
A_adapter_t | Long Y-adaptor (top) | AATGATACGGCGACCACCGAGATCTACACTCTTTCCCTACACGACGCTCTTCCGATC*T | 3'-Phosphorothioate bond | [1] | |
A_adapter_b | Long Y-adaptor (bottom) | GATCGGAAGAGCGGTTCAGCAGGAATGCCGAGACCGATCTCGTATGCCGTCTTCTGCTTG | None | [1] | |
PE_t_adapter | Regular Y-adaptor (top) | ACACTCTTTCCCTACACGACGCTCTTCCGATC*T | 3'-Phosphorothioate bond | [2] | |
PE_b_adapter | Regular Y-adaptor (bottom) | /5Phos/GATCGGAAGAGCGGTTCAGCAGGAATGCCGAG | 5'-phosphorylation | [2] | |
PCR_F | Sol. Amp. (Forward) | AATGATACGGCGACCACCGAGATCTACACTCTTTCCCTACACGACGCTCTTCCGATC*T | 3'-Phosphorothioate bond | [2] | |
PCR_R | Sol. Amp. (Reverse) | CAAGCAGAAGACGGCATACGAGATCGGTCTCGGCATTCCTGCTGAACCGCTCTTCCGATC*T | 3'-Phosphorothioate bond | [2] | |
Syb_FP5 | Sol. Quant. (Forward) | ATGATACGGCGACCACCGAG | None | [2] | |
Syb_RP7 | Sol. Quant. (Reverse) | CAAGCAGAAGACGGCATACGAG | None | [2] | |
PE_t_N2 | Y-adaptor for MmeI ligation (top) | ACACTCTTTCCCTACACGACGCTCTTCCGATCTN*N | 3'-Phosphorothioate bond | ||
PE_N2 | Y-adaptor for MmeI ligation (top) | ACACTCTTTCCCTACACGACGCTCTTCCGATC*NN | 3'-Phosphorothioate bond | ||
PE_b_A | Y-adaptor for MmeI ligation (bottom) | /5Phos/AGATCGGAAGAGCGGTTCAGCAGGAATGCCGAG | 5'-phosphorylation | ||
PE_t_Met | bis-seq(Forward) | A/iMe-dC/A/iMe-dC/T/iMe-dC/TTT/iMe-dC//iMe-dC//iMe-dC/TA/iMe-dC/ACGACG/iMe-dC/T/iMe-dC/TT/iMe-dC/CGAT/iMe-dC/*T | 3-phosphorothiate, 5'-Methyl | ||
PE_b_Met | bis-seq (Reverse) | /5Phos/GATCGGAAGAGCGGTT/iMe-dC/AG/iMe-dC/AGGAATG/iMe-dC/CGAG | 5'-phosphorylated, 5'-Methyl | ||
RC_PE_t_adapter | RC_Regular Y-adaptor with Biotin (top) | /5Biosg/TTTTTTTAGATCGGAAGAGCGTCGTGTAGGGAAAGAGTGT | 5'-Biotin | ||
RC_PE_b_adapter | RC_Regular Y-adaptor with Biotin (bottom) | /5Biosg/TTTTTTTCTCGGCATTCCTGCTGAACCGCTCTTCCGATC | 5'-Biotin | ||
Small RNA Sequencing Primer | Solexa-TSS-Seq | CGACAGGTTCAGAGTTCTACAGTCCGACGATC | |||
- [1] Amplification-free Illumina sequencing-library preparation facilatates improved mapping and assembly of GC biased genomes
- [1] supp
- [2] Improved protocols for the Illumina GA
Multiplexing Reverse PCR primers
- Primer list and assigned managers File:ZhangLab MultiplexingPrimers BarcodeInfo384 12212011.xls
Library Free Multiplexing Primers
Name | Nick-name | Sequence (5\'->3\') | Sequence Modification | Reference |
Left Probe Linker, No barcode (forward) | AmpF6.3Sol | AATGATACGGCGACCACCGACACTCTCAGATGTTATCGAGGTCCGAC | None | None |
Right Probe Linker, Barcode 1: CGTGAT (reverse) | AmpR6.3Ind1 | CAAGCAGAAGACGGCATACGAGATCGTGATGCTAGGAACGATGAGCCTCCAAC | None | None |
Right Probe Linker, Barcode 2: ACATCG (reverse) | AmpR6.3Ind2 | CAAGCAGAAGACGGCATACGAGATACATCGGCTAGGAACGATGAGCCTCCAAC | None | None |
Right Probe Linker, Barcode 3: GCCTAA (reverse) | AmpR6.3Ind3 | CAAGCAGAAGACGGCATACGAGATGCCTAAGCTAGGAACGATGAGCCTCCAAC | None | None |
Right Probe Linker, Barcode 4: TGGTCA (reverse) | AmpR6.3Ind4 | CAAGCAGAAGACGGCATACGAGATTGGTCAGCTAGGAACGATGAGCCTCCAAC | None | None |
Right Probe Linker, Barcode 5: CACTGT (reverse) | AmpR6.3Ind5 | CAAGCAGAAGACGGCATACGAGATCACTGTGCTAGGAACGATGAGCCTCCAAC | None | None |
Right Probe Linker, Barcode 6: ATTGGC (reverse) | AmpR6.3Ind6 | CAAGCAGAAGACGGCATACGAGATATTGGCGCTAGGAACGATGAGCCTCCAAC | None | None |
Right Probe Linker, Barcode 7: GATCTG (reverse) | AmpR6.3Ind7 | CAAGCAGAAGACGGCATACGAGATGATCTGGCTAGGAACGATGAGCCTCCAAC | None | None |
Right Probe Linker, Barcode 8: TCAAGT (reverse) | AmpR6.3Ind8 | CAAGCAGAAGACGGCATACGAGATTCAAGTGCTAGGAACGATGAGCCTCCAAC | None | None |
Right Probe Linker, Barcode 9: CTGATC (reverse) | AmpR6.3Ind9 | CAAGCAGAAGACGGCATACGAGATCTGATCGCTAGGAACGATGAGCCTCCAAC | None | None |
Right Probe Linker, Barcode 10: AAGCTA (reverse) | AmpR6.3Ind10 | CAAGCAGAAGACGGCATACGAGATAAGCTAGCTAGGAACGATGAGCCTCCAAC | None | None |
Right Probe Linker, Barcode 11: GTAGCC (reverse) | AmpR6.3Ind11 | CAAGCAGAAGACGGCATACGAGATGTAGCCGCTAGGAACGATGAGCCTCCAAC | None | None |
Right Probe Linker, Barcode 12: TACAAG (reverse) | AmpR6.3Ind12 | CAAGCAGAAGACGGCATACGAGATTACAAGGCTAGGAACGATGAGCCTCCAAC | None | None |
Forward sequencing primer | SolSeqV6.3.2 | ACCGACACTCTCAGATGTTATCGAGGTCCGAC | None | None |
Reverse sequencing primer | SolSeqV6.3.2r | GCTAGGAACGATGAGCCTCCAAC | None | None |
Index read primer | AmpR6.3IndSeq | GTTGGAGGCTCATCGTTCCTAGC | None | None |
Forward extension primer | AmpF6.3SolExt | AATGATACGGCGACCACCGAGATCTACACCACTCTCAGATGTTATCGA | None | None |
Forward PCR primer | AmpF6.4Sol | AATGATACGGCGACCACCGAGATCTACACCACTCTCAGATGTTATCGAGGTCCGAC | None | None |
Forward sequencing primer | SolSeqV6.3.3 | TACACCACTCTCAGATGTTATCGAGGTCCGAC | None | None |
- The current version of sequencing primers used for Illumina’s PE flowcell
- SolSeq6.3.3 (read1) : (5’-TACACCACTCTCAGATGTTATCGAGGTCCGAC -3’)
- SolSeqV6.3.2r (read2) : (5’-GCTAGGAACGATGAGCCTCCAAC-3’)
- AmpR6.3IndSeq (Index read primer) : (5’-GTTGGAGGCTCATCGTTCCTAGC-3’)
- LibFreeIndSeqRev (Index read primer after 2nd strand synthesis) : (5’-CAAGCAGAAGACGGCATACGAGAT-3’)
- 48-plex primers designed by Dinh.
- Media:Library-free_BarcodeInfo384_designedbyDinh_June16_2011-edit.xls
Multiplex primers for the N2-adaptor
Sequence ID | Sequence | |
PCR_R.N2Ind1 | CAAGCAGAAGACGGCATACGAGATCGTGATCTCGGCATTCCTGCTGAACCGCTCTT | |
PCR_R.N2Ind2 | CAAGCAGAAGACGGCATACGAGATACATCGCTCGGCATTCCTGCTGAACCGCTCTT | |
PCR_R.N2Ind3 | CAAGCAGAAGACGGCATACGAGATGCCTAACTCGGCATTCCTGCTGAACCGCTCTT | |
PCR_R.N2Ind4 | CAAGCAGAAGACGGCATACGAGATTGGTCACTCGGCATTCCTGCTGAACCGCTCTT | |
PCR_R.N2Ind5 | CAAGCAGAAGACGGCATACGAGATCACTGTCTCGGCATTCCTGCTGAACCGCTCTT | |
PCR_R.N2Ind6 | CAAGCAGAAGACGGCATACGAGATATTGGCCTCGGCATTCCTGCTGAACCGCTCTT | |
PCR_R.N2Ind7 | CAAGCAGAAGACGGCATACGAGATGATCTGCTCGGCATTCCTGCTGAACCGCTCTT | |
PCR_R.N2Ind8 | CAAGCAGAAGACGGCATACGAGATTCAAGTCTCGGCATTCCTGCTGAACCGCTCTT | |
PCR_R.N2Ind9 | CAAGCAGAAGACGGCATACGAGATCTGATCCTCGGCATTCCTGCTGAACCGCTCTT | |
PCR_R.N2Ind10 | CAAGCAGAAGACGGCATACGAGATAAGCTACTCGGCATTCCTGCTGAACCGCTCTT | |
PCR_R.N2Ind11 | CAAGCAGAAGACGGCATACGAGATGTAGCCCTCGGCATTCCTGCTGAACCGCTCTT | |
PCR_R.N2Ind12 | CAAGCAGAAGACGGCATACGAGATTACAAGCTCGGCATTCCTGCTGAACCGCTCTT | |
PCR_Fs | AATGATACGGCGACCACCGAGATCTACACTCTTTCCCTACACGACGCTCTTC | |
N2IndSeq | AAGAGCGGTTCAGCAGGAATGCCGAG | |
N2RevSeq(WRONG design) | CTCGGCATTCCTGCTGAACCGCTCTT | |
N2RevSeq2 | CTCGGCATTCCTGCTGAACCGCTCTTCCGATCT | |
N2FwdSeq2H (for overloaded flowcells only) | CACTCTTTCCCTACACGACGCTCTTCCGATCTH | |
N2RevSeq2H (for overloaded flowcells only) | TCGGCATTCCTGCTGAACCGCTCTTCCGATCTH |
Nextera Read Primer
Sequence ID | Sequence | |
Nextera Read 1 Primer | 5'-GCCTCCCTCGCGCCATCAGAGATGTGTATAAGAGACAG-3' | |
Nextera Read 2 Primer | 5'-GCCTTGCCAGCCCGCTCAGAGATGTGTATAAGAGACAG-3' | |
Nextera Index Read Primer | 5'-CTGTCTCTTATACACATCTCTGAGCGGGCTGGCAAGGCAGACCG-3' |
Nextera Primer
ID | Sequence | Length |
Nextera Primer 1* | AATGATACGGCGACCACCGA | 20nt |
Nextera Primer 2* | CAAGCAGAAGACGGCATACGA | 21nt |
Nextera Adaptor 1* | AATGATACGGCGACCACCGAGATCTACACGCCTCCCTCGCGCCATCAG | 48nt |
Nextera Adaptor 2 (minus bar code)* | CAAGCAGAAGACGGCATACGAGATCGGTCTGCCTTGCCAGCCCGCTCAG | 49nt |
Orange primer | CCTTGCCAGCCCGCTCAG | 18nt |
Blue primer | CCTCCCTCGCGCCATCAG | 18nt |
Short Orange primer | CCTTGCCAGCC | 11nt |
Short Blue primer | CCTCCCTCGCG | 11nt |
Primers for BSPP probes synthesized by Agilent
Sequence ID | Sequences | Length |
pAP1V61U | 5’-G*G*G*TCATATCGGTCACTGTU-3’ | 18 |
AP1V6 | 5\'- GTCATATCGGTCACTGTT -3\' | 18 |
AP2V6 | 5’-/5Phos/CACGGGTAGTGTGTATCCTG-3’ | 20 |
- * Indicates a phosphorothioate bond
Primers for LC Sciences probe experiment
Sequence ID | Sequnces | Length |
eMIP_CA1_F | TGCCTAGGACCGGATCAACT | 20 |
eMIP_CA1_R | GAGCTTCGGTTCACGCAATG | 20 |
CP-2-FA | GCACGATCCGACGGTAGTGT | 20 |
CP-2-RA | CCGTAATCGGGAAGCTGAAG | 20 |
CP-2-FA.IndSol | CAAGCAGAAGACGGCATACGAGATNNNNNNCGGTCTGCCATCCGACGGTAGTGT | 54 |
CP-2-RA.Sol | AATGATACGGCGACCACCGAGATCTACACGCCTATCGGGAAGCTGAAG | 48 |
Read 1: CP-2-SeqRead1.x | TACACGCCTATCGGGAAGCTGAAG | 24 |
Index Read: CP-2-IndSeq.x | ACACTACCGTCGGATGGCAGACCG | 24 |
Switch.CP-2-FA.Sol | AATGATACGGCGACCACCGAGATCTACACGCCTATCCGACGGTAGTGT | 48 |
Switch.CP-2-RA.IndSol | CAAGCAGAAGACGGCATACGAGATNNNNNNCGGTCTGCCATCGGGAAGCTGAAG | 54 |
Read 1 CP-2-SeqRead1.y | TACACGCCTATCCGACGGTAGTGT | 24 |
Index Read:CP-2-IndSeq.y | CTTCAGCTTCCCGATGGCAGACCG | 24 |
Primers for LC Sciences library-free protocol
Sequence ID | Sequences | Length |
CA-2-FA.Indx7Sol | 5’-CAAGCAGAAGACGGCATACGAGATGATCTGCGGTCTGCCATCCGACGGTAGTGT-3’ | 54 |
CA-2-FA.Indx45Sol | 5'-CAAGCAGAAGACGGCATACGAGATCGTAGTCGGTCTGCCATCCGACGGTAGTGT-3' | 54 |
CA-2-FA.Indx76Sol | 5'-CAAGCAGAAGACGGCATACGAGATAATAGGCGGTCTGCCATCCGACGGTAGTGT-3' | 54 |
CA-2-FA.Indx77Sol | 5'-CAAGCAGAAGACGGCATACGAGATACCCGTCGGTCTGCCATCCGACGGTAGTGT-3' | 54 |
CA-2-FA.Indx78Sol | 5'-CAAGCAGAAGACGGCATACGAGATAACACGCGGTCTGCCATCCGACGGTAGTGT-3' | 54 |
CA-2-RA.Sol | 5’-AATGATACGGCGACCACCGAGATCTACACGCCTATCGGGAAGCTGAAG-3’ | 48 |
CP-2-SeqRead1.x (Read1) | 5’-TACACGCCTATCGGGAAGCTGAAG-3’ | 24 |
CP-2-SeqRead2 | 5'-CGGTCTGCCATCCGACGGTAGTGT-3' | 24 |
CP-2-IndSeq.x (IndexRead) | 5’-ACACTACCGTCGGATGGCAGACCG-3’ | 24 |
Primers for totoRNAseq protocol
STRT-SEQ Read 1 v3
GATCTAAGCAGTGGTATCAACGCAGAGTACGGG 33nt Tm=71.2 (Kun's) correspond to TSO.r06 used for 5'end
totoRNAseq Read 1: [1]
CACCGAGATCTACACTCTTTCCCTACACGACGTT 34nt Tm=64(IDT) Tm=71.72(Kun's) correspond to TSO.idXX primers used for 5'end
totoRNAseq Read 1 v2:
CCACCGAGATCTACACTCTTTCCCTACACGACG 33nt Tm=64.5(IDT) Tm=72.04(Kun's) correspond to TSO.r02/TSO.r04 primers used for 5'end
RL.T7.IndSeq:
AAAAAAAAAAAAAAAAAAAAGCGGGCTGGCAAGG 34nt Tm=60.7(IDT) Tm=68.5(Kun's) correspond to RL.T7.id01-24 primers used for RT step (T20 based primers)
Primers for NEBNext Multiplex Oligos (updated May 08, 2013)
- Reference: [2]
NEBNext adaptor for Illumina: | 5´-/5Phos/GATCGGAAGAGCACACGTCTGAACTCCAGTC/ideoxyu/ACACTCTTTCCCT ACACGACGCTCTTCCGATC*T-3´ (loop adaptor, commercially purchased for ultra-low input ligation) |
Sequence ID | Sequences |
NEBNext Universal PCR Primer (PCR_F) | 5´-AATGATACGGCGACCACCGAGATCTACACTCTTTCCCTACACGACGCTCTTCCGATC-s-T-3´ |
NEBNext Index 1 | 5´-CAAGCAGAAGACGGCATACGAGATCGTGATGTGACTGGAGTTCAGACGTGTGCTCTTCCGATC-s-T-3´ |
NEBNext Index 2 | 5´-CAAGCAGAAGACGGCATACGAGATACATCGGTGACTGGAGTTCAGACGTGTGCTCTTCCGATC-s-T-3´ |
NEBNext Index 3 | 5´-CAAGCAGAAGACGGCATACGAGATGCCTAAGTGACTGGAGTTCAGACGTGTGCTCTTCCGATC-s-T-3´ |
NEBNext Index 4 | 5´-CAAGCAGAAGACGGCATACGAGATTGGTCAGTGACTGGAGTTCAGACGTGTGCTCTTCCGATC-s-T-3´ |
NEBNext Index 5 | 5´-CAAGCAGAAGACGGCATACGAGATCACTGTGTGACTGGAGTTCAGACGTGTGCTCTTCCGATC-s-T-3´ |
NEBNext Index 6 | 5´-CAAGCAGAAGACGGCATACGAGATATTGGCGTGACTGGAGTTCAGACGTGTGCTCTTCCGATC-s-T-3' |
NEBNext Index 7 | 5´-CAAGCAGAAGACGGCATACGAGATGATCTGGTGACTGGAGTTCAGACGTGTGCTCTTCCGATC-s-T-3´ |
NEBNext Index 8 | 5´-CAAGCAGAAGACGGCATACGAGATTCAAGTGTGACTGGAGTTCAGACGTGTGCTCTTCCGATC-s-T-3´ |
NEBNext Index 9 | 5´-CAAGCAGAAGACGGCATACGAGATCTGATCGTGACTGGAGTTCAGACGTGTGCTCTTCCGATC-s-T-3´ |
NEBNext Index 10 | 5´-CAAGCAGAAGACGGCATACGAGATAAGCTAGTGACTGGAGTTCAGACGTGTGCTCTTCCGATC-s-T-3´ |
NEBNext Index 11 | 5´-CAAGCAGAAGACGGCATACGAGATGTAGCCGTGACTGGAGTTCAGACGTGTGCTCTTCCGATC-s-T-3´ |
NEBNext Index 12 | 5´-CAAGCAGAAGACGGCATACGAGATTACAAGGTGACTGGAGTTCAGACGTGTGCTCTTCCGATC-s-T-3´ |
HP1 -> updated to HP6 (read1) | |
NEBNextRevSeq2 (read2) | 5' -GTGACTGGAGTTCAGACGTGTGCTCTTCCGATCT- 3' |
NEBNextIndSeq (index read) | 5' -GAAGAGCACACGTCTGAACTCCAGTCAC- 3' |
- -s- = phosphorothioate bond
- May 2013, more NEBNext Index primers (Index 9-32) in 96-well plate were ordered. The new primers do not have phosphorothioate bond at -s-T-3´ end as the ones from NEB kit.
- Updated ZhangLab_NEBNext_MultiplexingPrimers_BarcodeInfo_06142013
HiTSFLIP Sequencing Primer
- HiTSFLIP SEQ 1 5'GATCACGACGCTCTTCCGATCTCTACACTCTTTCCCTA
- Multiplexing Index Read Sequencing Primer 5' GATCGGAAGAGCACACGTCTGAACTCCAGTCAC
Adaptors/Primers for RRBS experiment, (updated Apr 24, 2014)
Barcoded adaptors
Name in Illumin TrueSeq | Sequences (5'-->3') |
TruSeq Universal Adapter | 5'- AATGATACGGCGACCACCGAGATCTACACTCTTTCCCTACACGACGCTCTTCCGATCT -3' |
TruSeq Adapter, Index XX | 5'- GATCGGAAGAGCACACGTCTGAACTCCAGTCACNNNNNNATCTCGTATGCCGTCTTCTGCTTG -3' |
PCR primers
TruS_F | 5'- AATGATACGGCGACCACCGAGATC -3' | Tm = 67.1 C |
TruS_R | 5'- CAAGCAGAAGACGGCATACGAGAT -3' | Tm = 65.5 C |
Read1: HP10
In dex read:HP12
Read2: HP11