Material | Vendor (Catalog Number) | |
PCR Plate | Fisher (E951020460) | |
8 Strip Flat Caps | Fisher (E0030124847) | |
Molecular Grade Water | Fisher (BP281910) | |
PCR 2X Master Mix | Promega (M7832) | |
Nested PCR and ASPE Primers | IDT | |
ExoSAP-IT | ThermoFisher (78201.1.ML) | |
dNTP Solution Set | NEB (N0446S) | |
Biotin-14-dATP | ThermoFisher (19524016) | |
NEB Taq Polymerase wih 10X Buffer | NEB (M0273S) | |
Streptavidin, R-Phycoerythrin Conjugate (SAPE) | Thermofisher (S-866) | |
xTAG 10X Buffer | Luminex (GR001C0060) | |
Drive Fluid | Luminex (40-50030) | |
Calibration Kit | Luminex (MPX-CAL-K25) | |
Performance Verification Kit | Luminex (MPX-PVER-K25) | |
Microspheres | Luminex | |
25 mL Reservoir | Fisher (14387070 | |
Amber 5 mL Tube | Axygen (MCT-500-X) | |
2 mL Tube | Eppendorf (022363352) | |
15 mL Tube | Eppendorf (0030122151) |
Bead Region | Catalog Number | Anti-TAG Sequence Bound to Each Microsphere (5'-3') | Complementary Primer TAG Sequence (5'-3') | Matching ospC Genotype | |
12 | MTAG-A012 | AGTAGAAAGTTGAAATTGATTATG | CATAATCAATTTCAACTTTCTACT | All | |
14 | MTAG-A014 | ATTGTGAAAGAAAGAGAAGAAATT | AATTTCTTCTCTTTCTTTCACAAT | A | |
15 | MTAG-A015 | GTTGTAAATTGTAGTAAAGAAGTA | TACTTCTTTACTACAATTTACAAC | B | |
18 | MTAG-A018 | GTAATTGAATTGAAAGATAAGTGT | ACACTTATCTTTCAATTCAATTAC | D | |
20 | MTAG-A020 | AAATTAGTTGAAAGTATGAGAAAG | CTTTCTCATACTTTCAACTAATTT | K | |
22 | MTAG-A022 | GATTGATATTTGAATGTTTGTTTG | CAAACAAACATTCAAATATCAATC | I/C | |
30 | MTAG-A030 | GTGTTATAGAAGTTAAATGTTAAG | CTTAACATTTAACTTCTATAACAC | M | |
36 | MTAG-A036 | TTGTGTAGTTAAGAGTTGTTTAAT | ATTAAACAACTCTTAACTACACAA | E | |
38 | MTAG-A038 | AGTAAGTGTTAGATAGTATTGAAT | ATTCAATACTATCTAACACTTACT | T | |
42 | MTAG-A042 | ATTTGTTATGATAAATGTGTAGTG | CACTACACATTTATCATAACAAAT | F | |
44 | MTAG-A044 | AATGTAAAGTAAAGAAAGTGATGA | TCATCACTTTCTTTACTTTACATT | G | |
46 | MTAG-A046 | GTGATTGAATAGTAGATTGTTTAA | TTAAACAATCTACTATTCAATCAC | H | |
48 | MTAG-A048 | TATGAATGTTATTGTGTGTTGATT | AATCAACACACAATAACATTCATA | I | |
52 | MTAG-A052 | GTAAGATTAGAAGTTAATGAAGAA | TTCTTCATTAACTTCTAATCTTAC | J | |
54 | MTAG-A054 | TAGAGAAAGAGAGAATTGTATTAA | TTAATACAATTCTCTCTTTCTCTA | L | |
56 | MTAG-A056 | AATTAGAAGTAAGTAGAGTTTAAG | CTTAAACTCTACTTACTTCTAATT | N | |
61 | MTAG-A061 | TATTAGAGAGAAATTGTAGAGATT | AATCTCTACAATTTCTCTCTAATA | O | |
63 | MTAG-A063 | TTTGTTGTTAAGTATGTGATTTAG | CTAAATCACATACTTAACAACAAA | U | |
65 | MTAG-A065 | TGAGTAAGTTTGTATGTTTAAGTA | TACTTAAACATACAAACTTACTCA | V | |
67 | MTAG-A067 | TTTGTGTGTTATTGTAATTGAGAT | ATCTCAATTACAATAACACACAAA | W | |
72 | MTAG-A072 | AATTGAGAAAGAGATAAATGATAG | CTATCATTTATCTCTTTCTCAATT | E/C |
Primer | Sequence (5'-3') | |
ospC ALL Tag 12 | CATAATCAATTTCAACTTTCTACTAGATTAGGCCCTTTAACAGACTCATC | |
ospC Type A Tag 14 | AATTTCTTCTCTTTCTTTCACAATATTGTGATTATTTTCGGTATCC | |
ospC Type B Tag 15 | TACTTCTTTACTACAATTTACAACCTCGTTGCGATTTGCTTCA | |
ospC Types E/C Tag 72 | CTATCATTTATCTCTTTCTCAATTTGCAAGTAAGGTCTCAACTT | |
ospC Types I/C Tag 22 | CAAACAAACATTCAAATATCAATCTCCGTTGTTATCTGCCTCATTATCT | |
ospC Type D Tag 18 | ACACTTATCTTTCAATTCAATTACATGATTATTTAGAGTGCCTAAAGCATTGTTTTGATC | |
ospC Type E Tag 36 | ATTAAACAACTCTTAACTACACAATGTGTTTTTACTCTGATTGGCCTCTAAACCATTATTGCC | |
ospC Type F Tag 42 | CACTACACATTTATCATAACAAATCGCCTGAACGCCTAAACCATTTGCATC | |
ospC Type G Tag 44 | TCATCACTTTCTTTACTTTACATTGGTGTTGTGATTCGCATCAG | |
ospC Type H Tag 46 | TTAAACAATCTACTATTCAATCACGCCCCCATCGTCACCCAAAGTGCCATTTTG | |
ospC Type I Tag 48 | AATCAACACACAATAACATTCATATTTGAAATTAAATATGCTCCTGA | |
ospC Type J Tag 52 | TTCTTCATTAACTTCTAATCTTACTCCGTTTTGACCCACTTCAGC | |
ospC Type K Tag 20 | CTTTCTCATACTTTCAACTAATTTCCCCGCTTCGACAGCTAAACCACCATTTTGTTG | |
ospC Type L Tag 54 | TTAATACAATTCTCTCTTTCTCTAATCGCTACCTAAAGTACCACCTGCTTC | |
ospC Type M Tag 30 | CTTAACATTTAACTTCTATAACACACCGGCATTTAAACCATTTTGGGCTATCAAA | |
ospC Type N Tag 56 | CTTAAACTCTACTTACTTCTAATTGTTTTGCACATCATCTAAACCATTATTATT | |
ospC Type O Tag 61 | AATCTCTACAATTTCTCTCTAATATTGGTTAACTAAGCCATTTGCC | |
ospC Type T Tag 38 | ATTCAATACTATCTAACACTTACTATGGCCTGCATCGACACT | |
ospC Type U Tag 63 | CTAAATCACATACTTAACAACAAACTGCCCTTGCAAGTCCTGT | |
ospC Type V Tag 65 | TACTTAAACATACAAACTTACTCAGAGCCGCTTGAGCAGTTAAACCATTTGCACC | |
ospC Type W Tag 67 | ATCTCAATTACAATAACACACAAATCGTTTCGATTTGCTTCTACACCC |
A | B | |
Primer | Sequence (5'-3') | |
ospC1F | ATGAAAAAGAATACATTAAGTGCA | |
ospC622RC | TTGGACTTTCTGCCACAACA |
A | B | |
Component | Volume (μL) | |
Promega PCR Master Mix (2X) | 12.5 | |
ospC1F (10 uM) | 0.5 | |
ospC622RC (10 uM) | 0.5 | |
Molecular grade water | 10.5 |
A | B | C | |
Temperature (°C) | Time | Cycles | |
95 | 2 minutes | 1 | |
95 | 30 seconds | 32 | |
49 | 30 seconds | 32 | |
72 | 50 seconds | 32 | |
72 | 10 minutes | 1 | |
4 | Hold | 1 |
A | B | |
Primer | Sequence (5'-3') | |
OC6(+)24 | AAAGAATACATTAAGTGCGATATT | |
OC602(-)22 | GGGCTTGTAAGCTCTTTAACTG |
A | B | |
Component | Volume μL | |
Promega PCR Master Mix (2X) | 12.5 | |
OC6(+)24 (10 μM) | 0.5 | |
OC602(-)22 (10 μM) | 0.5 | |
Molecular grade water | 10.0 |
A | B | C | |
Temperature (°C) | Time (minutes) | Cycles | |
37 | 15 | 1 | |
80 | 15 | 1 | |
4 | Hold | 1 |
A | B | |
Component | Volume (μL) | |
Molecular grade water | 9.75 | |
10X NEB PCR buffer | 2 | |
ASPE primer mix (500 nM) | 2 | |
ASPE dNTP mix (50 μM) | 1 | |
NEB Taq polymerase (5 U/μL) | 0.25 |
A | B | C | |
Temperature (°C) | Time | Cycles | |
95 | 2 minutes | 1 | |
95 | 30 seconds | 35 | |
56 | 30 seconds | 35 | |
68 | 30 seconds | 35 | |
68 | 5 minutes | 1 | |
4 | Hold | 1 |
A | B | C | |
Temperature (°C) | Time (minutes) | Cycles | |
96 | 2 | 1 | |
37 | 30 | 1 |