Hosuk:FISSEQ Primer information
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Primers[edit]
- FISSEQ_RT : RT primer for 1st Rolony fabrication
- /5Phos/TCTCGGGAACGCTGAAGANNNNNN
- FISSEQ_RCA : RCA primer for 1st Rolony fabrication
- TCTTCAGCGTTCCCGA*G*A
- FISSEQ_ppRCA : RCA primer for 2nd Rolony fab. from padlock probes
- GATATCGGGAAGCTGA*A*G
Padlock Probes[edit]
Structure[edit]
- Probe: H1 + Common seq. for 2nd RCA + Common Linker + decoding probe seq. + H2
- Common seq. for 2nd RCA : CTTCAGCTTCCCGATATC
- Common Linker : CGACGG
Padlock probes[edit]
- ppACTB
- H1:
- H2:
- /5Phos/CTGTGCTCGCGGGGCGCTTCAGCTTCCCGATATCCGACGGACGTATCGGTAGTCGCAACGCAGGCAAAGGCGAGGCT
- ppRAB7A
- H1:
- H2:
- /5Phos/GAAGCGAGAAGGTCCAAGTTCTGCTTCAGCTTCCCGATATCCGACGGGTCTTGCGTGCGATACGGAGTAGAGGAGACTAAACGGAGGACA
- ppMALAT1
- H1: TTTCTGCCTTTACTTATCAATTC
- H2: AATGGAGGTATGACATATAATCT
- /5Phos/TTTCTGCCTTTACTTATCAATTCCTTCAGCTTCCCGATATCCGACGGTCTACTTCGTCGCGTCAGACCAAATGGAGGTATGACATATAATCT
- ppMALAT1_NH2
- /5Phos/TTTCTGCCTTTACTTATCAATTCCTTCAGCTTCCCGATATCCGACGG/iUniAmM/TCTACTTCGTCGCGTCAGACCAAATGGAGGTATGACATATAATCT
- MALAT1_template
- /5AmMC6/GAATTGATAAGTAAAGGCAGAAAAGATTATATGTCATACCTCCAT
- ppCOL1A1 (dcProbe3-Cy3)
- /5Phos/CCTCAGCCTCCTGACTAGCTGCTTCAGCTTCCCGATATCCGACGGCCGTTTGATCGTTCGACCGAGACCGGGTTCATGCAATTCTCCTG
Detection Probes[edit]
- FISSEQ_Adpt : Cy3 labeled probe for 1st Rolony detection
- /5Cy3/TCTCGGGAACGCTGAAGA
- FISSEQ_AdptCy5 : Cy5 labeled probe for 1st Rolony detection
- /5Cy5/TCTCGGGAACGCTGAAGA
- ACTB_1stRolonyFISH : ATTO488 labeled probe for ACTB on 1st Rolony
- GGCAAAGGCGAGGCT/3ATTO488N/
- RAB7A_1stRolonyFISH : ATTO550 labeled probe for RAB7A on 1st Rolony
- GAGGAGACTAAACGGAGGACA/3ATTO550N/
- MALAT1_1stRolonyFISH : Cy3 labeled probe for MALAT1 on 1st Rolony
- /5Cy3/AATGGAGGTATGACATATAATCT
- Shifted FISSEQ_2ndRCAprimerDye : 2nd Rolony from all padlock probes
- GGATATCGGGAAGCTGAA/3ATTO488N/
- dcProbe0-Cy3 : for 2nd Rolony of ppACTB
- Cy3-ACGTATCGGTAGTCGCAACGCA
- dcProbe1-Cy3 : for 2nd Rolony of ppRAB7A
- Cy3-GTCTTGCGTGCGATACGGAGTA
- dcProbe2-Cy3 : for 2nd Rolony of ppMALAT1
- Cy3-TCTACTTCGTCGCGTCAGACCA
- dcProbe3-Cy3 : for 2nd Rolony
- Cy3-CCGTTTGATCGTTCGACCGAGA
- dcProbe4-Cy3 : for 2nd Rolony
- Cy3-CTACCATTCGCGTGCGATTCCA
- RT-dT50 : Cy3 labeled probe for mRNA detection
- /5Cy3/TTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTT
- RT-Adapt : Cy5 labeled probe for cDNA after RT
- /5Cy5/TCTTCAGCGTTCCCTAGA
- dcProbe2RevComp-Cy3 : ATTO488 labeled probe for detecting the reverse complementary sequence of dcProbe2 used for detecting ppMALAT1 and any other padlock probe before RCA with dcProbe2 in barcode
- TGGTCTGACGCGACGAAGTAGA/3ATTO488N/
Agi26k Probeset Barcode-to-dcProbe Key[edit]
[Position][Color] | Sequence | dcProbe |
[0][0] | ||
[0][1] | ATGTATCGCGCTCGATTGGCAG | dcProbe0-FAM |
[0][2] | ACGTATCGGTAGTCGCAACGCA | dcProbe0-Cy3 |
[0][3] | TACGCTACGGAGTACGCCACTC | |
[1][0] | ||
[1][1] | GTCTTGCGTGCGATACGGAGTA | dcProbe1-Cy3 |
[1][2] | CAACGGTATTCGGTCGTCATCC | dcProbe1-FAM |
[1][3] | ACTGGTTCGGGCGTACCTAACA | |
[2][0] | ||
[2][1] | TAGAACTTGCGCGGATACACGG | dcProbe2-FAM |
[2][2] | TCTACTTCGTCGCGTCAGACCA | dcProbe2-Cy3 |
[2][3] | GGACGAACGGTCGAGATTTACC | |
[3][0] | ||
[3][1] | CGAATTGTCCGCGCTCTACGAT | dcProbe3-FAM |
[3][2] | CCGTTTGATCGTTCGACCGAGA | dcProbe3-Cy3 |
[3][3] | AAACTGCGACCGTCGGCTTACT | |
[4][0] | ||
[4][1] | TCGGAATACGTCGTTGACTGCG | dcProbe4-FAM |
[4][2] | CTACCATTCGCGTGCGATTCCA | dcProbe4-Cy3 |
[4][3] | CCAGGGATCGGTCGAGTACGCT | |
[5][0] | ||
[5][1] | TGAGTGTCGCGCAACTTAGCGG | |
[5][2] | AACGTCTGCGTACCGGCTTAGC | |
[5][3] | ACATGCGATTAACCGCGACTGC | |
[6][0] | ||
[6][1] | TCACGCTTACGATCCCGCTATG | |
[6][2] | TTCGTAACCCGTGCGAAGTGCC | |
[6][3] | CCTCTCGTAGCGTGCGATGAGA |
- Position [7] (8th position if count first as 1) not used
Agi26k Probeset Barcode-to-dcProbe Key for dcProbes Ordered 5/14/14[edit]
[Position][Color] | Sequence | dcProbe |
[0][0] | ||
[0][1] | ATGTATCGCGCTCGATTGGCAG | dcProbe0-488 |
[0][2] | ACGTATCGGTAGTCGCAACGCA | dcProbe0-Cy3 |
[0][3] | TACGCTACGGAGTACGCCACTC | dcProbe0-Cy5 |
[1][0] | ||
[1][1] | GTCTTGCGTGCGATACGGAGTA | dcProbe1-488 |
[1][2] | CAACGGTATTCGGTCGTCATCC | dcProbe1-Cy3 |
[1][3] | ACTGGTTCGGGCGTACCTAACA | dcProbe1-Cy5 |
[2][0] | ||
[2][1] | TAGAACTTGCGCGGATACACGG | dcProbe2-488 |
[2][2] | TCTACTTCGTCGCGTCAGACCA | dcProbe2-Cy3 |
[2][3] | GGACGAACGGTCGAGATTTACC | dcProbe2-Cy5 |
[3][0] | ||
[3][1] | CGAATTGTCCGCGCTCTACGAT | dcProbe3-488 |
[3][2] | CCGTTTGATCGTTCGACCGAGA | dcProbe3-Cy3 |
[3][3] | AAACTGCGACCGTCGGCTTACT | dcProbe3-Cy5 |
[4][0] | ||
[4][1] | TCGGAATACGTCGTTGACTGCG | dcProbe4-488 |
[4][2] | CTACCATTCGCGTGCGATTCCA | dcProbe4-Cy3 |
[4][3] | CCAGGGATCGGTCGAGTACGCT | dcProbe4-Cy5 |
[5][0] | ||
[5][1] | TGAGTGTCGCGCAACTTAGCGG | dcProbe5-488 |
[5][2] | AACGTCTGCGTACCGGCTTAGC | dcProbe5-Cy3 |
[5][3] | ACATGCGATTAACCGCGACTGC | dcProbe5-Cy5 |
[6][0] | ||
[6][1] | TCACGCTTACGATCCCGCTATG | dcProbe6-488 |
[6][2] | TTCGTAACCCGTGCGAAGTGCC | dcProbe6-Cy3 |
[6][3] | CCTCTCGTAGCGTGCGATGAGA | dcProbe6-Cy5 |