Introduction
Results
Design of a multiplex PCR assay for SIV
Primer Set | Pool | Concentration (µM) | FWD sequence | REV sequence | Forward primer reference position | Reverse primer reference position |
---|---|---|---|---|---|---|
1 | 1 | 40 | TCCTGAGTACGGCTGAGTGAAG | CCTTCTTTGTTCTCCAACAGGCT | 1115–1137 | 1475–1498 |
2 | 2 | 30 | ATTAG G CTACG ACCCAACG G AA | AATTTCCTCCTCTG CCG CTAGA | 1363–1385 | 1711–1733 |
3 | 1 | 20 | ACCTAGTGGTG G AAACAGGAACA | GTCCTTGTTGTGGAGCTGGTTG | 1628–1651 | 1987–2009 |
4 | 2 | 20 | CCATCAAG CG G CTATG CAG ATT | GCTGCATCTGTCTGTTCTGCTC | 1893–1915 | 2246–2268 |
5 | 1 | 40 | TGGGGTTGCAAAAATGTGTCAGA | GCTTTCTTGGTCCCCTCTGTTG | 2114–2137 | 2476–2498 |
6 | 2 | 20 | GCCGGGACAGAAGGCTAGATTA | TTTAG CAG ATCCACAG CTG GGT | 2376–2398 | 2720–2742 |
7 | 1 | 30 | AATTTCCCCATG G CTCAAG TG C | TCCTATTCCTCCTACTATTTTTGGGGT | 2644–2666 | 2963–2990 |
8 | 2 | 10 | TG G ATACAG GGGCTG ATG ATTCT | CCAACTGACCATCCTTTTCCATCTTT | 2881–2904 | 3249–3275 |
9 | 1 | 20 | GTCGCCTTAAAGCCAGGAAAGG | GAGGTATGGAGAAATATGCATCACCT | 3135–3157 | 3465–3491 |
10 | 2 | 10 | TAGG AATACCACACCCTG CAGG | CCCTGTCATGTTCCAGGTCTGT | 3382–3404 | 3708–3730 |
11 | 1 | 40 | ACATG TG CTAG AACCCTTCAG G A | CAG TCCACTG AACTTCCTCTG TT | 3602–3625 | 3996–4019 |
12 | 2 | 10 | AGAGAGACCTGGACAGTG AATG A | TG TG CTAATAGTCTCACTCCATTG G T | 3852–3875 | 4213–4239 |
13 | 1 | 40 | GAGTCAGGACAATCAGTGGTCTTAT | ATCCTGCTTTCCCTTCTTTTGACTG | 4100–4125 | 4473–4498 |
14 | 2 | 10 | TCTCAACACCACCGCTAGTAAGA | TACCTTTGTGTGCTGGTACCCA | 4354–4377 | 4728–4750 |
15 | 1 | 15 | CCTACAG AATCAG AGAGCAGGCT | ATTTG CCTG CCCATGTATAG CC | 4632–4655 | 4949–4971 |
16 | 2 | 15 | CCCAG AATAGTG GCCAGACAGA | CAAAG G TG TG CTCTATCCCTG C | 4869–4891 | 5202–5224 |
17 | 1 | 30 | ATGGTG CTAACTTTG CTTCG CA | GGTCCCTTCCACAGTTGATCTCT | 5134–5156 | 5497–5520 |
18 | 2 | 30 | AGGGG ATATG ACTCCAGCAG AA | CCATCCG ACCTTAAAATG GGGC | 5357–5379 | 5742–5764 |
19 | 1 | 25 | AG AGGTGG ATAG CAGTTCCCAC | CTTCTCCCG CTG TAAAG CAAG G | 5609–5631 | 5959–5981 |
20 | 2 | 50 | GGACAGATGTAACACCAAACTATGCA | TTCCCAAG ACCTTTG CCAAACC | 5885–5911 | 6229–6251 |
21 | 1 | 70 | GGAGAAGAGACAATAGGAGAGGCC | CATCCCATGGTTCCCTTTGTGG | 6107–6131 | 6458–6480 |
22 | 2 | 10 | TCCCCCTCCAG G ACTAG CATAA | TG CTTCTAG AGGGCGG TATAG C | 6385–6407 | 6704–6726 |
23 | 1 | 30 | TCATG CATTTCAG AGGCGG ATG | GCATTCCTCCAAGCTGGTACAC | 6615–6637 | 6972–6994 |
24 | 2 | 10 | TTGGG AATCAG CTG CTTATCG C | TGTCAATCCCCATCTATCTGTCTCA | 6870–6892 | 7231–7256 |
25 | 1 | 25 | AG TCACAG AACAGG CAATAG AGGA | TCCCTTGTTCACATACCAAATCTGC | 7102–7126 | 7451–7476 |
26 | 2 | 40 | AG CTGTAAATTCAACATGACAG GGTT | GTCCTATTATCCCTACCATGCCAGT | 7358–7384 | 7749–7774 |
27 | 1 | 20 | GGTGGTCTCTTCATGCACAAGG | TGGGATGTTTGACAATGGTCTGC | 7633–7655 | 7955–7978 |
28 | 2 | 20 | AAAGCAGGCATGGTGTTGGTTT | TGGAGTTACACGTGAGGTCTCC | 7876–7898 | 8243–8265 |
29 | 1 | 20 | CCAG AAG CCAAAG GAACAG CAT | TTGCGAGAAAACCCAAGAACCC | 8113–8135 | 8471–8493 |
30 | 2 | 20 | AG ATG TG AAG AG GTACACTACTG G T | CTAAACGCACATCCCCAAGCAT | 8389–8414 | 8700–8722 |
31 | 1 | 15 | TAG G GTCACTG CCATCG AG AAG | TCCAAGAAGCAAGGTCAAACCA | 8632–8654 | 8921–8943 |
32 | 2 | 40 | AGCTGGG ATG TGTTTGG CAATT | GCCAAGTCAAGAGGCGTATCAG | 8879–8901 | 9206–9228 |
33 | 1 | 100 | GGCAAAGAAAGAGACGGTGGAG | AAGAGAGTGAGCTCAAGCCCTT | 9101–9123 | 9495–9517 |
34 | 2 | 40 | CTGGAGATCTGCG ACAG AGACT | GTAATAAATCCCTTCCAGTCCCCC | 9370–9392 | 9734–9758 |
35 | 1 | 40 | TTGGTAG G G GTATCAGTG AGGC | CCATGCTAGAACCTCTCCCCAA | 9621–9643 | 9971–9993 |
36 | 2 | 40 | CAGATGAGGCACAGGAGG ATG A | ACATCAAGAAAGTGGGCGTTCC | 9880–9902 | 10,205–10,227 |
37 | 1 | 40 | AGGG ACTTTCCACAAG GGG ATG | TCGGTTTCCCAAAGCAGAAAGG | 10,131–10,153 | 10,516–10,538 |
Detection of false positives in clonal SIVmac239
Detection of genome-wide variants using multiplex SIV sequencing
Position | SIVmac239 | SIVmac239-24x | Amplicon |
---|---|---|---|
1316 | T | C | 1 |
1481 | A | G | 2 |
1510 | G | A | 2 |
2467 | C | T | 6 |
2723 | A | G | 7 |
2850 | C | T | 7 |
2860 | G | A | 7 |
3721 | C | T | 11 |
4260 | G | A | 13 |
4945 | C | T | 15 |
5815 | C | A | 19 |
6199 | A | G | 20 |
6199 | A | G | 21 |
6639 | T | C | 22 |
6639 | T | C | 23 |
6923 | T | G | 23 |
6923 | T | G | 24 |
6925 | T | C | 23 |
6925 | T | C | 24 |
7058 | G | A | 24 |
8390 | G | A | 29 |
8750 | A | G | 31 |
8850 | A | G | 31 |
9110 | A | G | 32 |
9176 | A | G | 32 |
9181 | C | T | 32 |
9642 | C | T | 34 |
9934 | C | A | 35 |
9934 | C | A | 36 |