Table 3.
A comparison between targeted genome sequencing (TG-Seq) and gel electrophoresis (GE) to detect BYMV, PSbMV, PEBV and CMV amplicons generated in a multiplex PCR (mPCR) reaction.
| Library | Amplicons Targeted by mPCR a |
Raw Reads | No. of Reads after QC (%) |
Amplicons Detected by TG-Seq b |
Amplicons of BYMV, PSbMV, PEBV and CMV, Detected by GE |
|---|---|---|---|---|---|
| 1 | BYMV (NIb2, CPF3, HcProF2) | 3,754,078 | 97.74% | NIb (1,208,788), CP (1,503,144), HcPro (949,413) | CP, HcPro |
| 2 | BYMV (HcProF2, NIb3, CPF3) | 3,704,546 | 98.07% | HcPro (1,748,342), NIb (22,057), CP (1,839,520) | CP, HcPro |
| 3 | BYMV (NIb2, CP3, HcProF1) | 3,563,538 | 98.04% | NIb (1,487,879), CP (1,091,068), HcPro (906,692) | HcPro, NIb, CP |
| 4 | BYMV (CP, HcProF1, HcProF2) | 3,523,172 | 97.85% | CP (880,456), HcPro (2,490,101) | CP, HcPro |
| 5 | PSbMV (CP, NIb, HcPro, CI) | 4,568,980 | 98.71% | CP (967,475), NIb (1,005,493), HcPro (1,121,760), CI (1,403,227) |
CP, NIb, HcPro, CI |
| 6 | PEBV (12K, 14K, 30K, 201K) | 4,110,734 | 98.46% | 12K (706,420), 14K (979,796), 30K (1,371,956), 201K (978,813) |
12K, 14K, 30K, 201K |
| 7 | PEBV (12K, 14K, 30K, 201K) | 3,923,838 | 98.50% | 12K (515,527), 14K (900,724), 30K (153,998), 201K (899,718) | 12K, 14K, 30K, 201K |
| 8 | CMV (RNA1, RNA2, RNA3) | 3,457,376 | 98.44% | RNA1 (318,290), RNA2 (1,293,807), RNA3 (260,700) | RNA1, RNA2 |
| 9 | CMV (RNA1), PEBV (201K), PSbMV (CP) |
3,257,938 | 98.21% | RNA1 (1,299,800), 201K (1,145,237), CP (732,277) |
RNA1, 201K, CP |
| 10 | CMV(RNA3), PEBV (201K2), PSbMV (HcPro), BYMV (CP3) |
3,318,404 | 98.37% | RNA3 (207), 201K (1,561,718), HcPro (419,226, CP (1,248,550) |
201K, HcPro, CP3 |
| 11 | CMV (RNA1), PEBV (201K), PSbMV (CP), BYMV (HcPro) |
2,210,396 | 95.24% | RNA1 (703,928), 201K (8929), CP (5348), HcPro (1,057,739) |
RNA1, HcPro |
| 12 | CMV (RNA1), PEBV (201K), PSbMV (CP), BYMV (HcPro) | 2,514,042 | 98.54% | RNA1 (735,687), 201K (701,502), CP (645,571), HcPro (739,571) | RNA1, 201K, CP, HcPro |
This data was generated using the 16 primers designed in Table 2. The amplicon open reading frames (ORFs) targeted by mPCR are the BYMV and PSbMV each amplified using primer pairs designed from the NIb, CP, HcPro and CI (Libraries 1–5). PEBV multiplex reactions (library 7), targeting the 12K, 14K, 30K, and 201K ORFs of the PEBV genome. CMV multiplex reaction involved the three RNA1-3 components (library 8). Series of mPCR reactions to detect three to four plant viruses Libraries 9–12. a = Corresponding to virus name and specific primers name (s) listed in Table 2 and used to amplify the ORFs shown in the column named “Amplicons detected by TG-Seqb”, b = figures in parenthesis are the number of reads mapping to each genome region of interest.