1. Supplemental Fig. S1 Ihsan et al., 2015 Supplemental Fig. S1 Sequence analysis of Solanaceous MAGO NASHI genes. ClustalW multiple alignments of nucleotide sequences of the MAGO1 and MAGO2 genes classes of Withania somnifera (WsMAGO1 and WsMAGO2), Tubocapsicum anomalum (TaMAGO1 and TaMAGO2) and Physalis floridana (PfMAGO1 and PfMAGO2) in Bio-Edit program. Nucleotides in shaded blocks show conservation. Sequence duplication is enclosed by a solid blocked line.

2. Supplemental Fig. S2 Ihsan et al., 2015 Supplemental Fig. S2 Sequence analysis of Solanaceous MAGO NASHI proteins. ClustalW multiple alignments of amino acid sequences of the Solanaceous MAGO NASHI proteins and various other plants, animals, nematode and human generated in Bio-Edit program. Letter “L” in blue color and “R” in red color denote amino acids Leucine and Arginine, respectively. Two solid black line blocks enclose 3 amino acids (LRY) duplication. Arginine (R), Serine (S) and Glutamic acid (E) mutations are enclosed in solid block as SNPs. A consensus sequence is also given at the bottom.

3. Supplemental Fig. S3 Ihsan et al., 2015 Supplemental Fig. S3 WsMAGO2 and PfMAGO2 have divergent nucleotide sequences. ClustalW nucleotide alignment of WsMAGO2 and PfMAGO2 in Bio-Edit. Twelve nucleotides are different. Scale and consensus sequences are given.

4. Supplemental Fig. S4 Ihsan et al., 2015 Supplemental Fig. S4 WsMAGO2 and PfMAGO2 have identical amino acid sequences. ClustalW amino acid alignment of WsMAGO2 and PfMAGO2 in Bio-Edit. All the amino acids are identical. Scale and consensus sequences are given.

5. Supplemental Fig. S5 Generation of WsMAGO2 RNAi transgenic plants of Withania somnifera. a Photographs showing different stages of production of transgenic plants of W. somnifera. b Callus induction and subsequent multiple shoots regeneration from nodal explants of W. somnifera on MS media supplemented with 1mg/l kinetin and 2.5mg/l 6-benzylaminopurine, and 20 mg/l hygromycin for selection of transformed shoots for the WsMAGO2-RNAi knockout plants. c Sub-culturing of regenerated plantlets along with callus for further shoot multiplication. d root induction in plantlets on MS selection media fortified with indolebutyric acid (3mg/l). e Rooted transgenic plantlets. f Hardening of rooted transgenic plants in the humus soil-pearlite under controlled conditions. Supplemental Fig. S5 Ihsan et al., 2015

6. Supplemental Fig. S6 WsMAGO2 RNAi plants of Withania exhibit alterations in leaf development and calyx inflation. a Demonstration of leaf size variations of Ko106 and Ko112 knockdown lines along with wild type control plants. b Estimation of reduction in calyx inflation of transgenic Ko106 and Ko112 knockdown lines and control plants. Supplemental Fig. S6 Ihsan et al., 2015 a b

7. Supplemental Fig. S7 Ihsan et al., 2015 Supplemental Fig. S7 Amplification of WsMAGO2 in 14 transgenic plants for RNA estimation. RT-PCR analysis of WsMAGO2 knockdown W. somnifera plants along with wild type is shown. 18SrRNA served as an internal control for relative expression.

8. Supplemental Fig. S8 Ihsan et al., 2015 WsMAGO1 PfMAGO1 WsMAGO1 PfMAGO1 WsMAGO1 PfMAGO1 WsMAGO1 PfMAGO1 WsMAGO1 PfMAGO1 WsMAGO1 PfMAGO1 WsMAGO1 PfMAGO1 WsMAGO1 PfMAGO1 WsMAGO1 PfMAGO1 WsMAGO1 PfMAGO1 WsMAGO1 PfMAGO1 WsMAGO1 PfMAGO1 WsMAGO1 PfMAGO1 WsMAGO1 PfMAGO1 WsMAGO1 PfMAGO1 WsMAGO1 PfMAGO1 WsMAGO1 PfMAGO1 Supplemental Fig. S8 ClustalW nucleotide alignment of WsMAGO1 and PfMAGO1 promoters in Bio-Edit. Solid backward arrow indicates putative transcription initiation site. Anther and pollen specific motifs are highlighted in red and blue colors, and enclosed in a box.

9. Supplemental Fig. S9 Ihsan et al., 2014 Supplemental Fig. S9 ClustalW nucleotide alignment of WsMAGO2 and PfMAGO2 promoters in Bio-Edit. Solid backward arrow indicates putative transcription initiation site. Anther and pollen specific motifs are highlighted in red and blue colors, and enclosed in a box.

10. Supplemental Fig. S10 Ihsan et al., 2015 Supplemental Fig. S10 Pursued scheme of making WsMAGOs promoter:YFP/GUS constructs. A total of 6 constructs (2 for WsMAGO1promoter and 4 for WsMAGO2 promoter) 3 each for YFP and GUS reporters using pXCG-mYFP (upper panel) pGVT-bar as back bone vectors were generated, respectively. ∆ symbolizes deletion variant of WsMAGO2 promoter.