Supplemental Table 1: Oligonucleotide Primers Used in This Study

Supplemental Table 1: Oligonucleotide primers used in this study

Gene / Size / Forward (5'--->3') / Reverse (5'--->3')
PMT1 / 794 / ggcacacatgcttgtgacaaaagtaag / agttaactgctcacttcgtccctg
PMT2 / 733 / cgaggagtaaacagatgcgaagaaag / gagtaatatgcgaagttactatcc
PMT3 / 828 / tcgtctaatatgtatttttgcccatagac / ctcagtgcttagtcaaacaccatcatc
PMT4 / 932 / aggagtaaacagatgcgaagtaaga / agaataatatgcgaagttactggtc
A622 / 500 / tgtaatttcccgaggaaggattcaac / aaaagtagcgtaaccccataactc
700 / gtaacttgtcttatgttccagatcac / tgacaactttgtctctaggaggggt
881 / gcgatggaaatcccaaaggtaatgac / agatgaagtgccgatcaacatagtg
ADC1 / 828 / gacaaattctgccgctgcttcccatt / tgcttggtcctgagcttagccc
ADC2 / 659 / gacaaattctgccgctgcttccattg / tgcttggtcctgagcttagccc
MPO1 / 868 / ggtgtcatataaattgaaagtgaag / catatttaagccacaagattcaaggac
MPO2 / 802 / ggcccttctagtcactaactactgt / gacaacaacttgtgcaattgcacac
BBLa / 1037 / aaactgctactggagctgttac / gacgggagatatcaccagtcgag
BBLb / 1512 / agactcattctattacggcagggtc / ataaataattccccaatttcctcct
BBLd / 742 / aaggaatcatgctggtaatag / tactaggagagctcatggagatcgat
QPT2 / 655 / gtgtgattaagtttcttggaaatag / tgaaatactgaaaccaactcatcaatg
CYP82E4 / 1149 / tggaattatgcccatcctaca / cattagtggttgcacctgagg
811 / gatgagatgtgtgcatacttg / ccaaattagaaaaactcgtactg
CYP82E4v2 / 939 / attgtaggactagtaacccttacac / gaggcacaaagaattctcatc
615 / gagtagagggattgtttccg / gtacaatcaagataaaacatctaagg
CYP82E10 / 1148 / gtgatagtttgattcccaagtgc / ctcccaaagttagattagtccg
1086 / aggtcgcgctgattcttg / agatgaatacccatctatctaggagt
SAMS / 1386 / gttgatggaattctaatgttttgac / gctcatttgtgacaccttcatcatg
SPDS / 967 / tgggattgaggattctgcaaaga / tgccataagacactgctaatacatg
ERF189 / 1000 / cgtactactccattctaaaaagaatga / ctaaccacagccttgcaccttcacct
ERF115 / 681 / cagttatagcagcatttgtaggttc / taatgcaaccttctctttctctgag
JAZ1 / 990 / acagagggcaatttactagtgaccgagc / cggtctgtaatagctaacaccctatctag
1075 / cagttattctggtttcaagatttcagctc / gatcacatagcaataactttaccagct
COI1 / 2468 / tggtgaagctgtaggtttaacttgg / tctcacttctagcttttgcaggctagga
ORC1 / 620 / aaccctagtaagtgaaagtgaaag / cgatggctggggtgagcgcgtacggct
MYC1b / 1102 / ggatcataacaggtgtagaaattcgag / ttttcggtgttcttgctcagca
MYC2a / 1560 / cgatccgatatccgatccgatccaaac / ttccataccgagtattacttccatc
JAP1 / 683 / gaagactgggaagctccattaacgag / aatcactacacagaacccagttgg
JAT1 / 1631 / gatggtttcttgtaggtcatactag / ttgacatcactgatgtctcgagct
JAT2 / 1890 / tgagagattttagttcggtaataatta / catattgacatcactgatgtctcaagtg
NUP1 / 1156 / gcacgaggaaaatccaaaccttc / tcaaacagattcagtatgtgtcgtaa
NUP2 / 718 / gcacgagggaaaagcatttcagcc / ttgaatctctaacaccaacgtagc
NtMATE1 / 1142 / gaagttgagcagcccttgttgat / taactatgtagttttaattgccactg
NtMATE2 / 1394 / gaagttgagcagcccttgttggc / aagtgaatccgcaacgcgacatttgt
Sanger sequencing
PMT2intF / cacacatgtactaataaaattagggt
MPO1intF / atagtagtgttaattgatatgtaa
MYC2aintF / cagatctgaattgatttgtctg
RRTPCR
PMT / attggaccaagatcgagtc / attactgcagaattctcctac
ERF189 / gcagcttcgactgcagcttcct / ctcctcggactcggagcacttc
JAT1 / aatttcgaacacttcgatgg / tacccctaaattcgaacgcc
EF1α / aagcccatggttgttgagac / gtcaacgttcttgataacac

Supplemental Table 2: EMS induced mutations identified in TILLING population in different genes.

Gene / Position / Ref. base / Coverage / Total bases in alignment / Library / Location /Conseq
PMT1 / 346 / C / 11041 / A:10 / C:10971 / G:5 / T:55 / N:0 / 7 / 5'UTR
6583 / A:4 / C:6550 / G:1 / T:28 / N:0 / 9
A622B / 482 / G / 5556 / A:87 / C:4 / G:5453 / T:12 / N:0 / 3 / R109K
9942 / A:105 / C:7 / G:9811 / T:19 / N:0 / 15
ADC1 / 877 / C / 57720 / A:28 / C:57600 / G:16 / T:76 / N:0 / 3 / H79Q
68937 / A:39 / C:68784 / G:18 / T:95 / N:1 / 4
10010 / A:9 / C:9979 / G:3 / T:19 / N:0 / 14
ADC1 / 1167 / C / 10743 / A:18 / C:10671 / G:5 / T:49 / N:0 / 1 / A176V
17755 / A:39 / C:17649 / G:9 / T:57 / N:1 / 15
ADC2 / 414 / C / 10911 / A:27 / C:10825 / G:4 / T:55 / N:0 / 1 / A177V
33576 / A:33 / C:33490 / G:7 / T:46 / N:0 / 4
17700 / A:31 / C:17588 / G:9 / T:72 / N:0 / 15
MPO2 / 438 / G / 28075 / A:144 / C:15 / G:27867 / T:49 / N:0 / 3 / promoter
17955 / A:88 / C:5 / G:17843 / T:19 / N:0 / 13
BBLA / 104 / C / 29071 / A:19 / C:28924 / G:2 / T:126 / N:0 / 4 / 5'UTR
2227 / A:1 / C:2196 / G:1 / T:28 / N:1 / 9
BBLD / 342 / G / 3358 / A:72 / C:1 / G:3284 / T:1 / N:0 / 5 / R170Q
9234 / A:98 / C:3 / G:9127 / T:6 / N:0 / 15
CYP82E4_EX1 / 755 / G / 24723 / A:141 / C:6 / G:24553 / T:23 / N:0 / 3 / M224I
16470 / A:94 / C:3 / G:16357 / T:16 / N:0 / 11
Cyp82E4Ex2 / 390 / C / 42648 / A:46 / C:42499 / G:12 / T:91 / N:0 / 3 / Synonymous
50357 / A:49 / C:50221 / G:7 / T:78 / N:2 / 4
18344 / A:16 / C:18297 / G:6 / T:25 / N:0 / 9
Cyp82E4Ex2 / 458 / C / 15928 / A:16 / C:15889 / G:2 / T:21 / N:0 / 1 / A435V
50179 / A:39 / C:50046 / G:12 / T:82 / N:0 / 4
21529 / A:13 / C:21481 / G:6 / T:29 / N:0 / 14
CYP82E4v2_EX1 / 522 / G / 32021 / A:178 / C:6 / G:31814 / T:23 / N:0 / 4 / Synonymous
19045 / A:168 / C:4 / G:18854 / T:19 / N:0 / 14
Cyp82E10Ex1 / 165 / C / 54090 / A:53 / C:53940 / G:8 / T:89 / N:0 / 3 / P42L
42015 / A:57 / C:41863 / G:14 / T:81 / N:0 / 4
8332 / A:9 / C:8299 / G:3 / T:21 / N:0 / 13
8372 / A:12 / C:8336 / G:6 / T:18 / N:0 / 14
Cyp82E10Ex1 / 349 / C / 18341 / A:19 / C:17600 / G:7 / T:715 / N:0 / 7 / Synonymous
19930 / A:11 / C:19387 / G:7 / T:524 / N:1 / 13
Cyp82E10Ex2 / 697 / C / 13519 / A:16 / C:13388 / G:6 / T:109 / N:0 / 1 / A410V
32706 / A:38 / C:32427 / G:5 / T:236 / N:0 / 3
15931 / A:17 / C:15797 / G:5 / T:111 / N:1 / 12
19685 / A:19 / C:19512 / G:5 / T:149 / N:0 / 13
SAMS / 317 / C / 36074 / A:44 / C:35967 / G:5 / T:58 / N:0 / 3 / Not determined
21416 / A:24 / C:21352 / G:11 / T:29 / N:0 / 13
SPDS / 203 / C / 30693 / A:36 / C:29884 / G:9 / T:764 / N:0 / 3 / Intron
30236 / A:33 / C:29497 / G:10 / T:696 / N:0 / 4
9061 / A:11 / C:9023 / G:5 / T:20 / N:2 / 13
9582 / A:5 / C:9549 / G:5 / T:22 / N:1 / 15
SPDS / 529 / G / 24254 / A:471 / C:3 / G:23756 / T:23 / N:1 / 4 / Intron
14505 / A:61 / C:1 / G:14436 / T:7 / N:0 / 16
SPDS / 784 / G / 8282 / A:113 / C:3 / G:8159 / T:7 / N:0 / 3 / R119H
7613 / A:277 / C:2 / G:7327 / T:7 / N:0 / 7
5167 / A:87 / C:1 / G:5077 / T:2 / N:0 / 11
6474 / A:206 / C:2 / G:6263 / T:3 / N:0 / 16
ORC / 127 / C / 4072 / A:0 / C:4021 / G:1 / T:50 / N:0 / 1 / Not determined
4624 / A:2 / C:4555 / G:1 / T:66 / N:0 / 14
ORC / 436 / G / 25446 / A:333 / C:10 / G:25094 / T:9 / N:0 / 4 / not determined
12691 / A:159 / C:6 / G:12515 / T:11 / N:0 / 13
MYC1B / 858 / G / 10340 / A:74 / C:4 / G:10246 / T:16 / N:0 / 3 / V76I
11157 / A:45 / C:3 / G:11090 / T:19 / N:0 / 13
MYC2A / 237 / C / 10286 / A:11 / C:10075 / G:2 / T:198 / N:0 / 3 / promoter
7485 / A:9 / C:7337 / G:3 / T:136 / N:0 / 5
8214 / A:17 / C:8105 / G:2 / T:89 / N:1 / 12
8698 / A:5 / C:8542 / G:8 / T:143 / N:0 / 16
JAT1 / 1224 / C / 14811 / A:5 / C:14608 / G:1 / T:197 / N:0 / 8 / P153S
14266 / A:6 / C:14070 / G:0 / T:190 / N:0 / 16
JAT1 / 1435 / C / 10345 / A:10 / C:10247 / G:1 / T:87 / N:0 / 7 / S223F
9887 / A:5 / C:9773 / G:0 / T:109 / N:0 / 16
JAT2 / 191 / C / 5527 / A:2 / C:5379 / G:2 / T:144 / N:0 / 1 / promoter
20918 / A:22 / C:20566 / G:5 / T:325 / N:0 / 4
5795 / A:5 / C:5593 / G:2 / T:195 / N:0 / 10
8497 / A:10 / C:8277 / G:7 / T:202 / N:1 / 13
JAT2 / 633 / C / 14494 / A:21 / C:14242 / G:1 / T:230 / N:0 / 1 / Intron
22132 / A:15 / C:21953 / G:11 / T:152 / N:1 / 15
NUP1 / 408 / G / 10641 / A:43 / C:5 / G:10578 / T:15 / N:0 / 5 / D19N
15642 / A:235 / C:2 / G:15370 / T:35 / N:0 / 15
NUP1 / 443 / C / 12379 / A:12 / C:12053 / G:4 / T:310 / N:0 / 4 / Synonymous
8521 / A:15 / C:8424 / G:3 / T:79 / N:0 / 12
12757 / A:13 / C:12349 / G:2 / T:392 / N:1 / 13
NUP1 / 1807 / C / 15459 / A:17 / C:15371 / G:4 / T:67 / N:0 / 7 / Intron
11117 / A:11 / C:11042 / G:2 / T:61 / N:1 / 9
MATE1 / 384 / C / 19871 / A:16 / C:19812 / G:5 / T:38 / N:0 / 3 / Intron
18252 / A:19 / C:18050 / G:3 / T:180 / N:0 / 8
14126 / A:13 / C:13984 / G:8 / T:121 / N:0 / 12
MATE1 / 912 / C / 8864 / A:8 / C:8795 / G:2 / T:59 / N:0 / 4 / A194E
10709 / A:4 / C:10587 / G:2 / T:116 / N:0 / 14

Supplemental Table 3: Details of the tobacco (Nicotiana tabacum) germplasm accessions used in this study

S. No / Field code / PI Number / Variety/Cultivar / Origin / Type
1 / GP15 / PI405589 / Ethiopia / Oriental
2 / GP16 / PI405681 / ForchheimerHanica / Germany / Cigar filled
3 / GP24 / PI117037 / Amarillo / Peru / Primitive
4 / GP25 / PI119157 / Venezuela / Primitive
5 / GP27 / PI119452 / Caribe / Venezuela / Primitive
6 / GP28 / PI119454 / Venezuela / Primitive
7 / GP29 / PI119457 / Venezuela / Primitive
8 / GP38 / PI551318 / Speight G 28 / North Carolina / FCV
9 / GP44 / PI405682 / Geudertheimer III / Germany / FCV
10 / GP53 / PI418597 / Tan Yuh 1 / China / FCV
11 / GP55 / PI420186 / Lonibow / Ontario, Canada / FCV
12 / GP61 / PI431452 / N 209 / Japan / FCV
13 / GP65 / PI112355 / Virginia 312 / Columbia, U.S. / FCV
14 / GP77 / PI111459 / Columbia, U.S. / FCV
15 / GP78 / PI112127 / Bonanza / Columbia, U.S. / FCV
16 / GP83 / PI112162 / Cordoba / Columbia, U.S. / FCV
17 / GP84 / PI112165 / Cordoba / Columbia, U.S. / FCV
18 / GP86 / PI552752 / Smith 79x / North Carolina / FCV
19 / GP98 / PI552598 / Brown and Williams / North Carolina / FCV
20 / GP132 / PI112237 / Cash / Columbia, U.S. / FCV
21 / GP138 / PI113985 / Jaffna / North Carolina / FCV
22 / GP150 / PI119447 / Giron / Venezuela / FCV
23 / GP151 / PI119451 / Habano / Venezuela / FCV
24 / GP175 / PI118155 / Virginia / Venezuela / FCV
25 / GP180 / PI114564 / Mexico / FCV
26 / GP182 / PI114575 / Turco / Mexico / FCV
27 / GP194 / PI117669 / Goyano / Brazil / FCV
28 / GP216 / PI552381 / NC 75 / North Carolina / FCV
29 / GP219 / PI552328 / Paris Wrapper / Maryland / FCV
30 / GP241 / PI370291 / Ibsukhi ha / Japan / FCV
31 / GP245 / PI371613 / Lonibow / Taiwan / FCV
32 / GP255 / PI329207 / Immune 580 MS / Soviet Union / FCV
33 / GP261 / PI355075 / Sirogo / Australia / FCV
34 / GP268 / PI241765 / Chile / FCV
35 / GP277 / PI292204 / German virginia 230 / Poland / FCV
36 / GP279 / PI405547 / Turkey / FCV
37 / GP285 / PI405578 / Muscateler / Spain / FCV
38 / GP305 / PI405666 / V 20 / Poland / FCV
39 / GP312 / PI112203 / HabaneroLegtimo / Columbia / Primitive
40 / GP314 / PI110854 / Columbia / Cigar wrapper
41 / GP316 / PI109940 / Columbia / Cigar wrapper
42 / GP317 / PI405545 / South America / FCV
43 / GP318 / PI405544 / South America / FCV
44 / GP319 / PI405540 / Bright Yellow X Yenshu # 4 / Japan / FCV
45 / GP321 / PI405527 / Bolivia / FCV
46 / GP323 / PI113441 / Medellin / Columbia / Cigar filled
47 / GP324 / PI113590 / Punta de lanza / Columbia / Cigar filled
48 / GP327 / PI50840 / Nyanza / Burundi / Primitive
49 / GP332 / PI107475 / Maryland / Cigar wrapper
50 / GP357 / PI120513 / Turkey / FCV