Supplemental Table 1 the primers used in this study

Primer name / Sequence
ZmHKT1;1a-F / GTCGT CGTGCTGTATGTGCT
ZmHKT1;1a-R / TACGCACTGATGACCTCGAC
ZmHKT1;1b-F / CGACAATAGCACGAGCAAGA
ZmHKT1;1b-R / TCAGCACGTTGGATTATTGC
ZmGAPDH F / CCCTTCATCACCACGGACTAC
ZmGAPDH R / AACCTTCTTGGCACCACCCT
AY562132-F / GTAGCATTGTTGGTGGTGGTGTG
AY562132-R / ACCGTGGAGCAGTCAATGGAAG
AY554169-F / ACAGTCGTGAGCATTCCCAAC
AY554169-R / CCAAACCTTCTGTGCTACCTC
AY554170-F / CCGGAGTGAAGGGGATGG
AY554170-R / CAAGCAATGTGAATGGTATGTGAG
NtERD10B-F / CAATTTAGTGCAGGCCAGGC
NtERD10B-R / GGTCCATGGTGGCCAGGAAG
NtERD10C-F / GGGTAGCGCAAACGTGGAG
NtERD10C-R / CTTTTCCCTCAGCCTCGTGC
NtSOS1-F / CAAATGTTATCCCCCGAAAGC
NtSOS1-R / CGGAGAACCTGAGGAAATGTGA
NtNHX2-F / ACTCATCCCCATTGGTCCG
NtNHX2-R / AAGGAGTTCCACAAAAGCACGA
NtNHX4-F / CAAGAACTTCCGCACCCAC
NtNHX4-R / GCAGTATCAAACGCAGAGGACC
NtCAX3-F / CGGTTTGGCAATAATTGTCACAG
NtCAX3-R / CAACGATCATGCTTCAATCATCC

Supplemental Table 2 The stress-related genes

Gene name / GenBank accession number and major function
NtGPX / AB041518；encodes glutathione peroxidase；related to oxidative stress
NtSPS / AF194022；encodes Sucrose-6-phosphate synthase A；related to osmotic stress
NtERD10B / AB049336；encodes dehydrin；related to osmotic stress
NtRub-SS / X02353；encodes ribulose 1,5-bisphosphate carboxylase small
Subunit；involved in photosynthetic carbon assimilation/metabolism
NtAPX2 /

D85912；encodes cytosolicascorbate peroxidase； related to oxidative stress

NtCAX3 / -; vacuolar cation/proton exchanger 3；related to Ca2+ transport
NtRCA342 / U35111；encodes rubisco activase precursor；involved in photosynthetic carbon assimilation/metabolism
NtNHX4 / -；encodes sodium hydrogen exchanger 4；related to Na+ transport
NtAPX1 / AU15933；encodescytosolic ascorbate peroxidase；related to oxidative stress
ChlGaPA / M14417；encodes A-subunit of chloroplast glyceradehyde-3-P dehydrogenase；involved in photosynthetic carbon assimilation/metabolism
AY554170 / AY554170；encodes raffinose synthase family protein/seed imbibitions protein；related to osmotic stress

Supplemental Fig. 1 Vectors construction and molecular identification

(A)Schematic representation of the pCAMBIA3301-35S:ZmHKT1;1 constructs used to transform tobacco plants with the two transcripts of ZmHKT1;1 gene(ZmHKT 1;1a/b stands for the two transcripts);

(B)Verification of transgenic lines via PCR. Lane 1, H2O as blank control; Lane 2, WT plants as negative control; Lane 3, transgenic line 1-7 overexpressing ZmHKT 1;1a; Lane 4, transgenic line 1-13 overexpressing ZmHKT 1;1a; Lane 5, transgenic line 2-3 overexpressing ZmHKT 1;1b; Lane 6, transgenic line 2-8 overexpressing ZmHKT 1;1b;

(C) RT-PCR analysis of the expression level of ZmHKT 1;1 gene in T2 plants. Lane 1, H2O as blank control; Lane 2, WT plants as negative control; Lane 3, transgenic line 1-7 overexpressing ZmHKT 1;1a; Lane 4, transgenic line 1-13 overexpressing ZmHKT 1;1a; Lane 5, transgenic line 2-3 overexpressing ZmHKT 1;1b; Lane 6, transgenic line 2-8 overexpressing ZmHKT 1;1b;

Supplemental Fig.2Leaf disc assay of WT and transgenic lines (1-7,1-13,2-3 and 2-8) at different NaCl concentrations (0mM,200mM and 300mM) in T2 plants.

Supplemental Fig. 3Na+ and K+ contents in shoot(A and B) and root(C and D) of transgenic lines (1-7,1-13,2-3 and 2-8).

24-day-old WT and T2 transgenic plants were transferred into plastic containers containing improved Hoagland nutrient solution and pre-culture 3 days prior to receiving salt treatment of 0, 200 and 300mM NaCl. Shoot and root tissues were collected after 6 d of treatment and 3 d of recovery.Data are shown as the average ± S.E. of 15 plants. Experimental data was analyzed by t-test and the asterisks in columns mean significant difference from WT plants at *P<0.05 or **P<0.01 level.

Supplemental Fig. 4The relative expression changes of ZmHKT1;1a and ZmHKT1;1b in different tissues of maize.

The relative transcriptional level was analyzed using 2-ΔΔCt method, and the transcriptional level of ZmHKT1;1ain leaf was normalized as 1.00.

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