ars.els-cdn.com · Web viewSupporting information Fig ure S3 Automated ImageJ quantitative analysis of (a) In-situ accumulation of H 2 O 2 by DAB staining (b) (O 2 •─) anion radicals

Authors:

Krishna Kumar Raia,b, Nagendra Raib, Mohd Aamira, Deepika Tripathia and Shashi Pandey Raia*

Article Title:

Interactive role of salicylic acid and nitric oxide on transcriptional reprogramming for high temperature tolerance in Lablab purpureus

L.: Structural and functional insights using computational approaches

aCentre of Advance Study in Botany, Department of Botany, Institute of Science, Banaras Hindu University (BHU), Varanasi-221005 (Uttar

Pradesh), India

bIndian Institute of Vegetable Research, Post Box-01, P.O.-Jakhini (Shahanshahpur), Varanasi-221305 (Uttar Pradesh), India

Author for correspondence

*Email: [email protected]

aCentre of Advance Study in Botany, Department of Botany, Institute of Science, Banaras Hindu University (BHU), Varanasi-221005 (Uttar

Pradesh), India

mailto:[email protected]

Supporting information’s

Supporting information Figure S1 Enhanced fruit setting and number of pods in (B) SA 1 (0.5 mM), (C) SA 2 (1.0 mM), (D) SNP (0.5 mM),

(E) SA + SNP 1 (0.5 + 0.5 mM) and (F) SA + SNP 2 (1.0 + 0.5 mM) treated Lablab purpureus L plants (cv VRBSEM-15) compared to (A) NT:

non-treated HT stressed control plant under high temperature stress

Supporting information Figure S2 Aftereffect of different levels of salicylic acid (SA) and sodium nitroprusside (SNP) and their combinations

on (a) pollen germination and viability and (b) pollen tube growth in Lablab purpureus L (cv VRBSEM-15) under high temperature stress. NT:

non-treated HT stressed control, SA 1 (0.5 mM), SA 2 (1.0 mM), SNP (0.5 mM), SA + SNP 1 (0.5 + 0.5 mM) and SA + SNP 2 (1.0 + 0.5 mM).

Supporting information Figure S3 Automated ImageJ quantitative analysis of (a) In-situ accumulation of H2O2 by DAB staining (b) (O2•─)

anion radicals by NBT staining in Lablab purpureus L (cv VRBSEM-15) treated with different levels of salicylic acid (SA) and sodium nitroprusside (SNP) and their combinations under high temperature stress. NT: non-treated HT stressed control, SA 1 (0.5 mM), SA 2 (1.0 mM),

SNP (0.5 mM), SA + SNP 1 (0.5 + 0.5 mM) and SA + SNP 2 (1.0 + 0.5 mM). Mean (±SE) was calculated from three replicates for each treatment. Bars with distinct letters are significantly different at P ≤ 0.05 applying the DMRT test.

Supporting information Figure S4 Automated ImageJ quantitative analysis of changes in cross-sections of stem anatomy in Lablab purpureus L (cv VRBSEM-15) treated with different levels of salicylic acid (SA) and sodium nitroprusside (SNP) and their combinations under high

temperature stress. NT: non-treated HT stressed control, SA 1 (0.5 mM), SA 2 (1.0 mM), SNP (0.5 mM), SA + SNP 1 (0.5 + 0.5 mM) and SA + SNP 2 (1.0 + 0.5 mM). Mean (±SE) was calculated from three replicates for each treatment. Bars with distinct letters are significantly different at P ≤ 0.05 applying the DMRT test.

Supporting Information Figure S5. BLASTp results for the characterized protein (first hit JZ151191.1) as obtained through tBLASTn analysis of NCBI and searched across the annotated database of protein UniprotKB.

Supporting Information Figure S6. Phylogenetic tree showing the evolutionary origin and ancestral relationship in between the different HSF members based on uniprotKB.

Supporting Information Figure S7 Prediction of HSF02 proteins in Glycine max. genome as revealed through whole genome shortgun contig approach using tBLASTn programme of NCBI

Supporting information Figure S8 Prediction of potential genes in Glycine max genomic DNA

Table S1 Primers used for quantification of mRNA levels by qRT-PCR

Gene bank

(Accession number) GeneForward (5’- 3’) Reverse (5’- 3’)

JZ714621 ATP synthase CF1 alpha subunit AAAGCATCTTCTGTGGCTCAA GGCGCGAGGTATTGTAATGT

JZ714622 Chlorophyll a/b-binding protein CCTTTGGAGAACCTTGCTGA TCCGGGAACAAAGTTGGTAG

JZ714623 Lipid transfer protein CAATGCTGTGGAGGAGTCAA GCTGATTTCAAGCAGTTACAGG

JZ714624 Heat shock factor protein TTTTCAATATGGGTGCTGGTT CTGCCAAGGCTTTTATCTGG

JZ714625 Zinc finger protein CCACCGCTACTCCTTTCACT GCATGAAGATTGATCCATACGA

JZ714626 Thioredoxin h GAACAGCTCGAGAAGGGAAA GCCAAAATTGGAGCAATGAA

JZ714627 Late embryogenesis abundant protein ACGAGTGGAGGAGTGGTGTC CAGGATCAGGAACCCATGAA

JZ714628 Dicarboxylate transporter AAGTTGATTGGGTGGGTGGG CGTAGTGATTTGCGAGCGG

AJ222579 Elongation factor 1 alpha GACAACATGATTGAGAGGTCCACC GGCTCCTTCTCAATCTCCTTACC

Supporting Information Table S2 Functional annotation, accession ID and interacting score values for both first and second shell of interactors

that form mutual interactive associative network along with GmHSF02. The higher value of score indicate the more frequent interaction exist

between two associated proteins.

Node1 Node2 Node1 accession Node2 accession Node1 annotation Node2 annotation ScoreGLYMA01G01990.1 GLYMA02G47210.

1GLYMA01G01990.1 GLYMA02G47210.1 Uncharacterized protein Uncharacterized protein 0.768

GLYMA01G01990.1 GLYMA09G16690.1

GLYMA01G01990.1 GLYMA09G16690.1 Uncharacterized protein Uncharacterized protein 0.812

GLYMA01G01990.1 GLYMA09G24410.1


GLYMA01G01990.1 GLYMA12G07770.1


GLYMA01G01990.1 GLYMA14G01530.1


GLYMA01G01990.1 GLYMA14G40320.1


GLYMA01G01990.1 GLYMA16G29750.1


GLYMA01G01990.1 GLYMA17G37820.1


GLYMA01G01990.1 HSP90-1 GLYMA01G01990.1 GLYMA18G08220.1 Uncharacterized protein HSP90-1 0.768GLYMA01G01990.1 HSP90-2 GLYMA01G01990.1 GLYMA08G44590.1 Uncharacterized protein HSP90-2; Uncharacterized

protein0.768

GLYMA02G47210.1 GLYMA01G01990.1


GLYMA02G47210.1 GLYMA09G16690.1


GLYMA02G47210.1 GLYMA14G40320.1


GLYMA02G47210.1 GLYMA17G37820.1


GLYMA09G16690.1 GLYMA01G01990.1


GLYMA09G16690.1 GLYMA02G47210.1


GLYMA09G16690.1 GLYMA09G24410.1


GLYMA09G16690.1 GLYMA14G01530.1


GLYMA09G16690.1 GLYMA16G29750.1



protein0.936

GLYMA09G24410.1 GLYMA01G01990.1


GLYMA09G24410.1 GLYMA09G16690.1


GLYMA09G24410.1 GLYMA14G40320.1


GLYMA09G24410.1 GLYMA17G37820.1


GLYMA12G07770.1 GLYMA01G01990.1


GLYMA14G01530.1 GLYMA01G01990.1


GLYMA14G01530.1 GLYMA09G16690.1


GLYMA14G01530.1 GLYMA14G40320.1


GLYMA14G01530.1 GLYMA17G37820.1


GLYMA14G40320.1 GLYMA01G01990.1


GLYMA14G40320.1 GLYMA02G47210.1


GLYMA14G40320.1 GLYMA09G24410.1


GLYMA14G40320.1 GLYMA14G01530.1


GLYMA14G40320.1 GLYMA16G29750.1



protein0.933

GLYMA16G29750.1 GLYMA01G01990.1


GLYMA16G29750.1 GLYMA09G16690.1


GLYMA16G29750.1 GLYMA14G40320.1


GLYMA16G29750.1 GLYMA17G37820.1


GLYMA17G37820.1 GLYMA01G01990.1


GLYMA17G37820.1 GLYMA02G47210.1


GLYMA17G37820.1 GLYMA09G24410.1


GLYMA17G37820.1 GLYMA14G01530.1


GLYMA17G37820.1 GLYMA16G29750.1



protein0.933

HSP90-1 GLYMA01G01990.1

GLYMA18G08220.1 GLYMA01G01990.1 HSP90-1 Uncharacterized protein 0.768

HSP90-1 GLYMA09G16690.1


HSP90-1 GLYMA14G40320.1


HSP90-1 GLYMA17G37820.1


HSP90-2 GLYMA01G01990.1

GLYMA08G44590.1 GLYMA01G01990.1 HSP90-2; Uncharacterized protein

Uncharacterized protein 0.768

HSP90-2 GLYMA09G16690.1



HSP90-2 GLYMA14G40320.1



HSP90-2 GLYMA17G37820.1



Supporting Information Table S3 Functional annotation, accession ID and interacting score values for both first and second shell of interactors

that form mutual interactive associative network for lablab homologue GmHSF02

Node1 Node2 Node1 accession Node2 accession Node1 annotation Node2 annotation ScoreGLYMA01G01990.1

GLYMA09G16690.1 GLYMA01G01990.1

GLYMA09G16690.1 Uncharacterized protein Uncharacterized protein 0.812

GLYMA01G01990.1

GLYMA14G40320.1 GLYMA01G01990.1


GLYMA01G01990.1

GLYMA16G29750.1 GLYMA01G01990.1


GLYMA01G01990.1

GLYMA17G37820.1 GLYMA01G01990.1


GLYMA01G01990.1

HSP90-1 GLYMA01G01990.1

GLYMA18G08220.1 Uncharacterized protein HSP90-1 0.768

GLYMA06G00310.2

GLYMA09G16690.1 GLYMA06G00310.2


GLYMA06G00310.2

GLYMA14G40320.1 GLYMA06G00310.2


GLYMA06G00310.2

GLYMA16G29750.1 GLYMA06G00310.2


GLYMA06G00310.2

GLYMA17G37820.1 GLYMA06G00310.2


GLYMA06G00310.2

HSP90-1 GLYMA06G00310.2


GLYMA09G16690.1

GLYMA01G01990.1 GLYMA09G16690.1


GLYMA09G16690.1

GLYMA06G00310.2 GLYMA09G16690.1


GLYMA09G16690.1

GLYMA11G02341.1 GLYMA09G16690.1


GLYMA09G16690.1

GLYMA13G10700.1 GLYMA09G16690.1


GLYMA09G16690.1

GLYMA16G29450.1 GLYMA09G16690.1


GLYMA09G16690. GLYMA16G29750.1 GLYMA09G16690. GLYMA16G29750.1 Uncharacterized protein Uncharacterized protein 0.936

1 1GLYMA09G16690.1

GLYMA20G16070.1 GLYMA09G16690.1


GLYMA09G16690.1

HSP90-1 GLYMA09G16690.1


GLYMA11G02341.1

GLYMA09G16690.1 GLYMA11G02341.1


GLYMA11G02341.1

GLYMA14G40320.1 GLYMA11G02341.1


GLYMA11G02341.1

GLYMA16G29750.1 GLYMA11G02341.1


GLYMA11G02341.1

GLYMA17G37820.1 GLYMA11G02341.1


GLYMA11G02341.1

HSP90-1 GLYMA11G02341.1


GLYMA13G10700.1

GLYMA09G16690.1 GLYMA13G10700.1


GLYMA13G10700.1

GLYMA14G40320.1 GLYMA13G10700.1


GLYMA13G10700.1

GLYMA16G29750.1 GLYMA13G10700.1


GLYMA13G10700.1

GLYMA17G37820.1 GLYMA13G10700.1


GLYMA13G10700.1

GLYMA20G16070.1 GLYMA13G10700.1


GLYMA13G10700.1

HSP90-1 GLYMA13G10700.1


GLYMA14G40320.1

GLYMA01G01990.1 GLYMA14G40320.1


GLYMA14G40320.1

GLYMA06G00310.2 GLYMA14G40320.1


GLYMA14G40320.1

GLYMA11G02341.1 GLYMA14G40320.1


GLYMA14G40320.1

GLYMA13G10700.1 GLYMA14G40320.1


GLYMA14G40320.1

GLYMA16G29450.1 GLYMA14G40320.1



1 1GLYMA14G40320.1

GLYMA20G16070.1 GLYMA14G40320.1


GLYMA14G40320.1

HSP90-1 GLYMA14G40320.1


GLYMA16G29450.1

GLYMA09G16690.1 GLYMA16G29450.1


GLYMA16G29450.1

GLYMA14G40320.1 GLYMA16G29450.1


GLYMA16G29450.1

GLYMA16G29750.1 GLYMA16G29450.1


GLYMA16G29450.1

GLYMA17G37820.1 GLYMA16G29450.1


GLYMA16G29450.1

HSP90-1 GLYMA16G29450.1


GLYMA16G29750.1

GLYMA01G01990.1 GLYMA16G29750.1


GLYMA16G29750.1

GLYMA06G00310.2 GLYMA16G29750.1


GLYMA16G29750.1

GLYMA09G16690.1 GLYMA16G29750.1


GLYMA16G29750.1

GLYMA11G02341.1 GLYMA16G29750.1


GLYMA16G29750.1

GLYMA13G10700.1 GLYMA16G29750.1


GLYMA16G29750.1

GLYMA14G40320.1 GLYMA16G29750.1


GLYMA16G29750.1

GLYMA16G29450.1 GLYMA16G29750.1


GLYMA16G29750.1

GLYMA17G37820.1 GLYMA16G29750.1


GLYMA16G29750.1

GLYMA20G16070.1 GLYMA16G29750.1


GLYMA17G37820.1

GLYMA01G01990.1 GLYMA17G37820.1


GLYMA17G37820.1

GLYMA06G00310.2 GLYMA17G37820.1



1 1GLYMA17G37820.1

GLYMA13G10700.1 GLYMA17G37820.1


GLYMA17G37820.1

GLYMA16G29450.1 GLYMA17G37820.1


GLYMA17G37820.1

GLYMA16G29750.1 GLYMA17G37820.1


GLYMA17G37820.1

GLYMA20G16070.1 GLYMA17G37820.1


GLYMA17G37820.1

HSP90-1 GLYMA17G37820.1


GLYMA20G16070.1

GLYMA09G16690.1 GLYMA20G16070.1


GLYMA20G16070.1

GLYMA13G10700.1 GLYMA20G16070.1


GLYMA20G16070.1

GLYMA14G40320.1 GLYMA20G16070.1


GLYMA20G16070.1

GLYMA16G29750.1 GLYMA20G16070.1


GLYMA20G16070.1

GLYMA17G37820.1 GLYMA20G16070.1


GLYMA20G16070.1

HSP90-1 GLYMA20G16070.1


HSP90-1 GLYMA01G01990.1 GLYMA18G08220.1

GLYMA01G01990.1 HSP90-1 Uncharacterized protein 0.768













HSP90-1 GLYMA17G37820.1 GLYMA18G08220. GLYMA17G37820.1 HSP90-1 Uncharacterized protein 0.933

1HSP90-1 GLYMA20G16070.1 GLYMA18G08220.

1GLYMA20G16070.1 HSP90-1 Uncharacterized protein 0.876

Supporting Information Table S4 Mean minimum and maximum temperature, relative humidity, precipitation and source of irrigation during experimental period.

Temperature RH (%) Precipitation (mm)

Source of irrigation

Min. (ºC) Max. (ºC)7 days after sowing 32.2 43.3 79.3 10 Irrigation through handmade

water canal (twice a week)14 days after sowing 34.6 42.2 77.8 13 Irrigation through handmade






water canal (twice a week)Mean 36.1 42.2 81.8 23

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ars.els-cdn.com · Web viewSupporting information Fig ure S3 Automated ImageJ quantitative analysis of (a) In-situ accumulation of H 2 O 2 by DAB staining (b) (O 2 •─) anion radicals