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Origins of 229 wheat cultivars (lines) and their His and PFT genotypes
Detection of Fhb1 in 73 Yangmai and Ningmai cultivars (lines)
Specific primers used to amplify the open reading frames (ORF) of PFT, His, and HC
Subcellular localization of BnA7HSP70 and GFP fusion protein in Arabidopsis protoplasts.
(A) Subcellular localization of full-length GFP fused with green fluorescent protein (GFP), the cytoplasm showed a green fluorescent signal at 488 nm. (B) The mesophyll cells showed a red fluorescent signal at 580 nm. (C) A bright-field image of protoplast cell; (D) A bright-field image and the merged image are shown at the bottom. Scale bars, 10 µm.
BnA7HSP70 overexpressed (OE) plants confer tolerance under a restricted water regime and 20% PEG treatment
(A) For the fast soil drying treatment, wild type (WT) and BnA7HSP70 overexpressed (OE) lines were allowed to reach four to five weeks leaves stage of development when drought was rapidly induced by withholding irrigation for 10 days. The stress condition was prolonged until the leaves of wild type plants completely wilted. (B) For the fast soil drying treatment, wild type (WT) and BnA7HSP70 overexpressed (OE) lines were allowed to reach four to five weeks leaves stage of development when drought was rapidly induced by withholding irrigation for 15 days. The stress condition was prolonged until the leaves of wild type plants completely wilted. (C) After rewatering for 3 days, most wild type plants were unable to recover, while OE plants survived continued to grow. (D) 40-day-old WT and OE transgenic plants grown in nutrient solution. (E) For drought stress, 40-day-old WT and OE transgenic plants were transferred into nutrient solution containing 20% (w/v) PEG-6000 for 48 h. The concentration of PEG was maintained daily by changing the nutrient solution. (F) Leaf relative water content from WT and OE plants after 10 days, 15 days drought stress and 3 days rewater treatment. (G) Leaf relative water content from WT and OE plants after 20% PEG-6000 treatment for 48 h.
Changes of H₂O₂ content and MDA levels in wild plant and transgenic plant line under 20% PEG treatment.
(A) H₂O₂ content of seedlings at these days of the 20% PEG treatment; (B) MDA levels of seedlings at these days of the 20% PEG treatment. Data are shown as mean±SD of three independent measurements.
Antioxidant enzyme activities in wild plant and transgenic plants after treatment with 20% PEG
(A) SOD activity; (B) POD activity. Data are shown as means ±SD of three independent measurements.
Overexpression of BnA7HSP70 makes Brassica napus hold more water in soil pot without irrigation for 10 days
(A) Relative water content (RWC) of wild type and BnA7HSP70 overexpressing seedling (OE2, OE3, OE7, and OE8) leaves under drought condition. (B) Chlorophyll a and b concentrations were calculated as described in the materials and methods 1.5.3 and combined to give the total chlorophyll concentration, each of which is a mean of the samples taken from 6-8 leaf disks in each pool. Control: the wild and OE plants grew in the irrigated condition; Stress: four-week old wild and OE plants were subjected to progressive drought for seven days.
BnA7HSP70 overexpression delays drought-induced leaf senescence in OE lines confronted with stress
Drought was induced in wild type (WT) and OE transgenic plants (OE2, OE3, OE7, and OE8) at four-week old stage by withholding irrigation for 10 days. Control: normally irrigated plants. Stress: drought-stressed plants. Values are given as mean SD from three replicates. The experiment of senescence-associated genes BnLSC45 and BnLSC222 was induced by drought treatment. Total RNA was isolated from irrigated and drought-stressed mature leaves of wild plants and OE lines, and gene induction was monitored by quantitative RT-PCR using gene-specific primers.
BnA7HSP70 overexpression increases resistance against tunicamycin (TUN)-induced cell death
(A) Seeds and seedlings from overexpressed plants (OE) and untransformed wild-type (WT) plants were exposed to 2.5 µg mL^-1 or 5.0 µg mL^-1 tunicamycin. (B)-(C) Seedlings were monitored for the development of chlorosis and necrotic lesions, and cell viability were measured by the Evans blue dye method. Abs (600 nm) reflects the dead cell content. The values represent the average of three replicates (±SD).
Model for BnA7HSP70 expression on the mobilization of bZIP28 and upregulation of UPR genes
(A) In response to the stress, BnA7HSP70 is competed away by the accumulation of misfolded proteins and bZIP28 is proteolytically activated by Golgi-localized S1P or S2P to release bZIP28n, which relocates to the nucleus where it upregulates stress genes including BnBiP3 and BnCNX1; (B) When BnA7HSP70 is overexpressed, accumulated BnA7HSP70 is enough for association with bZIP28 and misfolded proteins. As a result, bZIP28 is detained in the ER even under stress conditions.
Performance of FHB resistance in the backcrossing progenies with or without Fhb1 gene from different donors, recurrent parent and controls using the floret-inoculation method
The number of diseased spikelets of backcrossing progenies with Fhb1 gene from donors Ningmai 9 (NM 9, A), Shengxuan 6 (SX 6, B), Jianyang 798 (JY 798, C), Jianyang 84 (JY 84, D), Sumai 3 (SM 3, E), and Ningmai 13 (NM 13, F) were obviously lower than the backcrossing progenies without Fhb1 gene. The recurrent parent Zhoumai 16 (ZM 16) and the highly susceptible control Annong 8455 (AN 8455) had more number of diseased spikelets than Huaimai 20 (HM 20), the moderately susceptible control (G) and Yangmai 158 (YM 158), the moderately resistant control (H). +: Fhb1 genotype; –: non-Fhb1 genotype; RP: recurrent parent; MS: moderately susceptible; HS: highly susceptible; MR: moderately resistant.
Analysis of variance (ANOVA) of Fusarium head blight index in 229 wheat cultivars
Structure, primer binding sites and three alleles of PFT gene
The first nucleotide of the initiation codon reads as +1
Amplification profiles of marker PFT-CAPS in partial Chinese cultivars
M: DL5000 DNA marker; 1-4: PFT-I cultivars, namely Sumai 3, Ningmai 9, Xinong 9871, and Xiaoyan 22; 5-8: PFT-II cultivars, namely Zhengmai 366, Lumai 21, Hengguan 35, and Ningchun 4. PCR products were digested with Dra I restriction endonuclease.
Alleles of His and HC genes for the cultivars with PFT-I and their FHB indexes
Amplification profiles of marker His-InDel in partial Chinese cultivars
M: DL5000 DNA marker; 1-4: His-I cultivars, namely Sumai 3, Ning 7840, Ningmai 9, and Ningmai 13; 5-11: non His-I cultivars, namely Xinong 9871, Xiaoyan 22, Zhengmai 9023, Yan 2415, Yangmai 158, Jimai 22, and Aikang 58.
Average FHB indexes of cultivars or lines with different PFT/His haplotypes
Comparison of number of diseased spikelets and disease index between backcrossing progenies with Fhb1 gene from different donors and recurrent parent, moderately susceptible control and the backcrossing progenies without Fhb1 gene
Comparisons of the number of diseased spikelets and disease index were carried out between all Fhb1 backcrossing progenies and the recurrent parent Zhoumai 16 (A), the moderately susceptible control Huaimai 20 (B) and all non-Fhb1 backcrossing progenies (C), and between non-Fhb1 and Fhb1 backcrossing progenies from the donor cultivars Ningmai 9 (D), Shengxuan 6 (E), Jianyang 798 (F), Jianyang 84 (G), Sumai 3 (H) and Ningmai 13 (I). NDS: number of diseased spikelets (floret-inoculation method); DI: disease index (natural infection nursery); ++: all Fhb1 backcrossing progenies; RP: recurrent parent; MS: moderately susceptible control; ––: all Fhb1 backcrossing progenies; +: Fhb1 backcrossing progenies; –: non-Fhb1 backcrossing progenies. * and ** above the error bars indicate significant difference at P < 0.05 and P < 0.01, respectively.
Comparison of number of diseased spikelets and disease index among the backcrossing progenies with Fhb1 gene from different donors

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