Variation in concentrations of highmolecularweight ...

Research Article Received: 5 October 2011

Revised: 25 December 2011

Accepted: 26 December 2011

Published online in Wiley Online Library: 6 February 2012

(wileyonlinelibrary.com) DOI 10.1002/jsfa.5607

Variation in concentrations of high-molecularweight glutenin subunits and macropolymers in wheat grains of a recombinant inbred lines population and in two contrasting eco-sites in China Xiangnan Li,a Jian Cai,a Fulai Liu,b Yun Bo,a Zengjun Qi,a Tingbo Dai,a Weixing Caoa and Dong Jianga∗ Abstract BACKGROUND: Concentrations of high-molecular-weight glutenin subunits and macropolymers in wheat grains are important indicators of grain quality, which are genetically determined and affected by environmental factors. The 6 VS·6AL translocation chromosome segment is reported to own high powdery mildew and yellow rust resistance genes of Pm21 and Yr26. This study investigated the variation in concentrations of high-molecular-weight glutenin subunits (HMW-GS) and glutenin macropolymer (GMP) in response to the 6 VS·6AL translocation segment and the two contrasting sites. RESULTS: Large variations in concentrations of HMW-GS and GMP were observed within lines containing different HMW-GS compositions and between the contrasting eco-sites. However, 6 VS·6AL chromosome translocation segment showed no significant effects on concentrations of HMW-GS and GMP. In addition, HMW-GS concentration was also found to be significantly correlated with the GMP concentration. CONCLUSION: Concentrations of HMW-GS and GMP are largely affected by the eco-sites and the composition of HMW-GS, whilst not by the presence of 6 VS·6AL chromosome segment translocation. The 6 VS·6AL translocation is suggested as potential donor for breeding wheat cultivars for high resistence to powdery mildew and yellow rust with less risk of undesirable effects on grain quality. c 2012 Society of Chemical Industry Supporting information may be found in the online version of this article. Keywords: glutenin subunits; protein component; 6 VS·6AL chromosome translocation segment; wheat (Triticum aestivum L.)

INTRODUCTION

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Haynaldia villosa L. Schur (syn. Dasypyrum villosum L. Candargy) (2n = 14, VV), a diploid allogamous wild Mediteranean grass species and a relative of bread wheat, is recognised as the tertiary gene pool for wheat improvement. Besides its high resistance to wheat diseases of powdery mildew,1 eyespot2 and rusts,3 Haynaldia villosa L. Schur is considered to carry genes expressing high grain protein concentration and strong gluten strength.4 Triticum aestivum L.–Haynaldia villosa L. Schur 6 VS·6AL translocation lines have been constructed and identified to contain genes of Pm21 and Yr26 which are highly resistant to powdery mildew and yellow rust.1 Hence, these lines have been extensively used in wheat breeding as disease-resistant donors. In China, several cultivars developed from 6 VS·6AL translocation lines have been released in many wheat production areas including Jiangsu, Henan and Hebei Provinces.5 These cultivars, such as Nannong 9918, Nannong 02Y393 and Shimai 14, are found to be highly resistant to powdery mildew.6

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The result of a test-cross has proved that the translocated chromosome 6 VS·6AL could be stably descended to offspring.7 However, other undesirable traits might be introduced into the offspring together with disease tolerance. Li et al.5 confirmed that many agronomic traits significantly differed between 6 VS·6AL translocation lines and their recurrent parents, including grain yield, grain number and weight per spike, and quality traits such as grain protein content, mixographic and farinographic

∗

Correspondence to: Dong Jiang, College of Agriculture, Nanjing Agricultural University, No.1 Weigang Road, Nanjing Jiangsu Province 210095, P.R. China. E-mail: [email protected]

a Key Laboratory of Crop Physiology and Ecology in Southern China, Ministry of Agriculture/Hi-Tech Key Laboratory of Information Agriculture of Jiangsu Province, Nanjing Agricultural University, P.R. China b University of Copenhagen, Faculty of Life Sciences, Department of Agriculture and Ecology, Høbakkegaard Allé 13, DK-2630 Taastrup, Denmark

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c 2012 Society of Chemical Industry

Eco-site affects wheat grain glutenin accumulation performances. However, using an F8 recombinant inbred lines (RIL) population derived from a Triticum aestivum L.–Haynaldia villosa L. Sxhur 6 VS·6AL translocation line 92R137 crossed with a Chinese wheat landrace Huixianhong, Wang et al.8 found that the translocation did not significantly affect grain protein content, flour yield, dough development time, area under extension and extensibility curve, and showed positive effects on maximum resistance to extension and stability, while it had negative effects on grain test weight, peak viscosity and softening degree of the dough. Thus, the effects of 6 VS·6AL translocation on wheat quality traits including protein composition still seem uncertain. Wheat grain protein is mainly composed of albumin, globulin, gliadin and glutenin. Of these components, glutenin macropolymers are the main determinant factor of flour baking quality9 and play important roles in dough development.10 Glutenin macropolymer (GMP) is mainly composed of oligomers of high-molecularweight glutenin subunits (HMW-GS) and low-molecular-weight glutenin subunits (LMW-GS),11 whereas the composition of HMWGS is closely correlated to the handling quality of wheat flour.12 It has been acknowledged that accumulation and concentration of GMP and HMW-GS in wheat grain are affected by both environmental variables such as extreme temperature,13 nitrogen14 and drought and water-logging,15 and by both genetic factors.16 However, little is known whether the 6 VS·6AL translocation regulates the concentrations of HMW-GS and GMP in wheat grain. In the present study, an F8 RIL population derived from the 6 VS·6AL translocation line 92R137 crossed with a Chinese landrace, Huixianhong, were grown in two contrasting ecological sites. Our purpose was to investigate the variation in concentrations of HMW-GS and GMP in wheat grain as related to 6 VS·6AL translocation and the growing sites. The results should help to better understand the values of 6 VS·6AL translocation in wheat breeding towards improved grain quality and disease resistance.

MATERIALS AND METHODS An F8 RIL population with 131 lines was constructed by the singleseed descendant from the cross of Huixianhong and 92R137. The former is an elite wheat landrace in the Huang-Huai-Hai plain, China, while the later was a typical 6 VS·6AL translocation line. The population was grown at both the Experimental Station of Nanjing Agricultural University, Nanjing (118◦ 42 E, 32◦ 30 N), Jiangsu Province, and the Experimental Station of Henan Agricultural University, Zhengzhou (113◦ 42 E, 34◦ 48 N), Henan Province, China, in the wheat growing season of 2005–2006. The former eco-site locates in the Yangtze River downstream plain, with a subtropical climate (average daily temperature 15.4 ◦ C, and average annual rainfall of 1200.0 mm), while the latter is in the Huang-Huai-Hai plain, with a warm temperate climate (average daily temperature 14.4 ◦ C, and average annual rainfall of 640.9 mm). Both regions are major wheat production areas, which account for more than 70% of the wheat production in China (Chinese Agriculture Annual Statistics, 2007). The field experiments were a randomised block design with three replicates for each line. Each line was planted into a three-row plot with a size of 1.5 × 0.75 m2 . The seedling density was 1.8 × 106 ha−1 with a row space of 25 cm. The plots were fertilised at both sites. Other field management was conducted following the local practices.

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by the sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) according to Khan et al.17 In brief, 40 mg grain sample was defatted with chloroform and then mixed with 1 mL of extraction buffer containing 62.5 mmol L−1 Tris-HCl (pH 6.8), 10% glycerol, 2% SDS and 5% β-mercaptoethanol. The mixture was incubated at room temperature for 30 min with continuous shaking, and then at 90 ◦ C for 5 min, followed by centrifugation at 8000 × g for 15 min. The supernatant was used for SDS-PAGE. The SDS-PAGE gel was 16 cm in both length and width, and 1 mm thick. The acrylamide concentration was 10% in the resolving gel and 4% in the stacking gel. Glutenin extract (25 µL) was loaded in each lane. After electrophoresis, the gel was stained with 0.05% Coomassie brilliant blue R250 for 24 h, and then destained in distilled water for 48 h. The destained gel was scanned using the VersaDoc Model 5000 Imaging Systems, using the Quantity 1-D Analysis Software (Bio-Rad Laboratories, Hercules, CA, USA) to give the preliminary content of each HMW-GS. Thereafter, each band was separately cut from the gel, and then placed in an Eppendorf tube. Depending on the intensity of each band, 500–1000 µL of 50% isopropyl alcohol containing 3% SDS was added to the tube and incubated at 37 ◦ C for 24 h to fully extract the dyestuff of Coomassie brilliant blue R250 conjugating with the subunit in the gel band. The concentration of the dyestuff extraction is completely dependent on the amount of HMW-GS in the corresponding band. The dyestuff extraction concentration was monitored at 595 nm with a UV-2401 Shimadzu spectrophotometer (Shimadzu Corp., Kyoto, Japan). During the preliminary electrophoresis, wheat cultivars of Chinese Spring and Marquis were used as standards to identify HMW-GS types of the selected lines. In the quantification procedure, 20, 30 and 40 µL of the standard protein (116 kDa; Sigma, St Louis, MO, USA) with given concentrations were separately loaded into three lanes on the same gel. The standard proteins loaded in the three lanes were quantified following the above protocol to obtain a standard curve. Concentration of each HMW-GS was then quantified based on the standard curve. Each sample was loaded into two lanes of the same gel, and the electrophoresis was performed at least twice for each sample. The average of the four lanes was taken as the mean concentration of each HMW-GS for the given sample. Glutenin macropolymer content GMP content was measured following the protocol of Weegels et al.9 In short, grain sample (50 mg) was suspended in 1 mL of 1.5% SDS solution and centrifuged at 15 500 × g for 30 min at 20 ◦ C. The nitrogen content of the sediment measured with the biuret reagent18 was taken as the GMP content. Statistical analysis All data were subjected to one-way ANOVA analysis using the SAS statistical analysis procedures (SAS Institute, Cary, NC, USA). Correlation analysis was conducted using datasets between content of HMW-GS and GMP, and the t-test was used to check 2 the significance. The χ 2 -test (χ 2 < χ0.05 ) was performed, which confirmed that the HMW-GS composition in grains of the offspring were evenly derived from their parents.

RESULTS Variation of the high-molecular-weight glutenin subunit composition in the recombinant inbred lines population and the subgroups Using NAU/xibao15902 as a co-dominant marker for identifying Pm21 located on 6 VS,19 52 lines from the 131 F8 RIL were confirmed


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Quantification of high-molecular-weight glutenin subunits The quantification of HMW-GS in wheat grain was performed following our previous protocol.14 HMW-GS were first separated

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the subunit/pair encoding loci were evenly transferred to the offspring. Eight HMW-GS combinations were detected in the RIL population (Table 1), including null, 7 + 8, 5 + 10 (GS1); null, 7 + 8, 5 + 12 (GS2); null, 7 + 9, 5 + 10 (GS3, the same as 92R137); null, 7 + 9, 5 + 12 (GS4); 1, 7 + 8, 5 + 10 (GS5); 1, 7 + 8, 5 + 12 (GS6, the same as Huixianhong); 1, 7 + 9, 5 + 10 (GS7) and 1, 7 + 9, 5 + 12 (GS8). It was noteworthy that the frequency of each HMW-GS combination did not vary greatly within the RIL population and within the subgroups. The frequencies of the two parent types, GS3 and GS6, were 12.21% and 13.75% in the whole RIL population. In the R subgroup, the frequency of GS5 was the highest (21.15%), followed by GS6 (15.38%) and GS3 (13.46%), and of GS1, GS2, GS4 and GS7 were the lowest (9.62%). The highest and the lowest frequency type was GS7 (16.46%) and GS5 (8.86%) in the S subgroup, respectively.

Subunit 1 5

2 7

7 9 10

8 12 CS

H

92R137

R

S

M

Figure 1. SDS-PAGE image of HMW-GS in wheat grain. Lane from the left to right is Chinese spring, Huixianhong, 92R137, R groups and S groups, Marquis, respectively. 1, 2, 5, 7, 8, 9, 10 and 12 on the left and right of figure indicates subunits 1, 2, 5, 7, 8, 9, 10 and 12, respectively.

to contain the 6 VS·6AL homozygous translocation and denoted as the R subgroup, while the remaining 79 lines did not carry the 6 VS·6AL and were denoted as the S subgroup. SDS-PAGE analysis confirmed that 92R137 contained HMW-GS subunits of null, 7 + 9 and 5 + 10, while Huixianhong contained subunits of 1, 7 + 8 and 5 + 12 (Fig. 1). SDS-PAGE analysis also revealed that the three pairs of HMW-GS encoded genes at the three complex loci of Glu-A1, Glu-B1 and Glu-D1 were segregated with a ratio of 1 : 1 in both the whole RIL population and the two subgroups (supplementary Table S1 and Table S2). In addition, no significant difference 2 ) in the frequency of each subunit/pair was observed (χ 2 < χ0.05 in the RIL population and in the two subgroups, indicating that

Table 1.

X Li et al.

Variation in concentrations of high-molecular-weight glutenin subunits and glutenin macropolymer A normal distribution of total HMW-GS concentration in the RIL population was found at both Nanjing and Zhengzhou sites (Fig. 2). The concentration among the whole population varied considerably with a large coefficient of variation of 33.61–43.39% (Table 2). In addition, variation in GMP concentration in the RIL population also showed a normal distribution at Nanjing and Zhengzhou sites (Fig. 3).

Line number and frequency of each HMW-GS combination in the recombinant inbred line (RIL) population and two subgroups RIL population

HMW-GS pair GS1a GS2 GS3 GS4 GS5 GS6 GS7 GS8 Total

R subgroup

S subgroup

Number

Frequency (%)

Number

Frequency (%)

Number

Frequency (%)

15 17 16 14 18 18 18 15 131

11.45 12.98 12.21 10.69 13.75 13.75 13.75 11.45 100

5 5 7 5 11 8 5 6 52

9.62 9.62 13.46 9.62 21.15 15.38 9.62 11.54 100

10 12 9 9 7 10 13 9 79

12.66 15.19 11.39 11.39 8.86 12.66 16.46 11.39 100

a GS1–GS9 indicates different HMW-GS combinations, i.e. GS1 − 7 + 8, 5 + 10; GS2 − 7 + 8, 5 + 12; GS3 − 7 + 9, 5 + 10; GS4 − 7 + 9, 5 + 12; GS5 − 1, 7 + 8, 5 + 10; GS6 − 1, 7 + 8, 5 + 12; GS7 − 1, 7 + 9, 5 + 10; GS8 − 1, 7 + 9, 5 + 12.

20

25 A

Nanjing

Zhengzhou

B

Number of lines

20

15

15 10 10 5

5 0

0 0

2

4

6

8

10

12

0

2

4

6

8

10

12

14

16

HMW-GS concentration (µg mg−1)

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Figure 2. Distribution of HMW-GS concentration in grains of the RIL population in Nanjing (A) and Zhengzhou (B).



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Eco-site affects wheat grain glutenin accumulation

Table 2.

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HMW-GS concentration in grains of the recombinant inbred line (RIL) population and two subgroups Glu-1A Population

Mean (µg·mg−1 )

Nanjing

Rb

0.73a

Zhengzhou

S W R S W

0.95b 0.87a 1.18a 1.38a 1.31a

Site

Glu-1B CV (%)c

Mean (µg·mg−1 )

53.45 48.81 51.92 53.99 38.73 43.06

2.61c

Glu-1D

2.46c 2.52b 3.63a 3.24b 3.41a

CV (%)

Mean (µg·mg−1 )

35.26 42.01 39.47 34.99 33.25 33.57

2.75c 2.57c 2.64b 3.95a 3.41b 3.63a

Total CV (%)

Mean (µg·mg−1 )

CV (%)

36.09 40.72 38.99 32.74 31.33 31.84

5.75c

33.61 43.39 39.91 34.20 35.88 34.93

5.51c 5.61b 8.20a 7.32b 7.62a

a

Different superscript letters in the same column indicate significant difference at P < 0.05 level. W, R and S indicate the whole population, the R- and S- subgroups, respectively. c CV, coefficient of variation. b

25

20

Number of lines

A

Nanjing

B

Zhengzhou 20

15

15 10 10 5

5

0

0 0

10

20

30

40

50

60

70 0

10

20

30

40

50

60

70

GMP concentration (µg mg−1) Figure 3. Distribution of GMP concentration in grains of the RIL population in Nanjing (A) and Zhengzhou (B).

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Correlation between concentrations of glutenin macropolymer and high-molecular-weight glutenin subunits There was a significantly positive correlation between GMP concentration and the concentrations of HMW-GS pairs of Glu-B1 and Glu-D1 (Fig. 4). The correlation coefficient between concentrations of GMP and HMW-GS pairs 7 + 8 (0.71) was the highest, followed by subunit pairs 5 + 12 (0.66), 5 + 10 (0.62), and 7 + 9 (0.55), and was the lowest for subunit 1 (0.20) in Nanjing. In Zhengzhou, GMP concentration had the highest correlation coefficient with the subunit pairs 7 + 8 (0.86), 5 + 12 (0.80) and 5 + 10 (0.80), followed by 7 + 9 (0.76), and the lowest with single subunit 1 (0.29).

DISCUSSION In the present study, recombination inbred lines derived from the cross of Huixianhong with 92R137 (carries the T6VS·6AL chromosome translocation segment) were grown in two ecosites. The lines were categorised into subgroups containing and non-containing the T6VS·6AL chromosome segment, and were identified as eight HMW-GS combinations. The analysis of the concentration of HMW-GS and GMP in grains revealed that variations in HMW-GS concentrations in the grains were more closely related to HMW-GS composition and the eco-sites, rather than the 6 VS·6AL translocation. A close relationship between concentrations of HMW-GS and GMP was also observed. HMW-GS are encoded by the polymorphic genes at the Glu-1 loci on the long arms of the group 1 chromosomes.16 Hexaploid



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The composition of HMW-GS significantly affected HMW-GS concentration in wheat grain (Table 3). Firstly, the presence of GluA1 had a significantly positive effect on HMW-GS concentration, as exemplified by much higher HMW-GS concentration in lines with subunit 1 than the counterpart lines without subunit 1. Secondly, HMW-GS concentration in lines owning subunit pair of 7 + 8 was higher than the counterpart lines owning subunit pair of 7 + 9. However, no clear tendency was observed in the variation of HMW-GS concentration between lines with a subunit pair of 5 + 10 and of 5 + 12. This indicated that subunit pairs of 7 + 8 usually cause a higher HMW-GS concentration than do pairs of 7 + 9, whilst the contribution to HMW-GS by subunit pairs 5 + 10 is similar to 5 + 12. The GMP concentration basically paralleled the HMW-GS concentration in the wheat grain. Comparison of the HMW-GS concentration between the S and the R subgroups revealed that T6VS·6AL translocation had no significant effect on grain HMW-GS concentration. However, the eco-sites significantly affected HMW-GS concentration of the same subgroup (Table 3). For instance, the mean HMW-GS concentration was 5.55 µg mg−1 in both R and S subgroups in Nanjing, whist was 7.66 µg mg−1 in the R subgroup and 7.36 µg mg−1 in the S subgroup in Zhengzhou. GMP concentration showed a similar tendency in responding to the 6 VS·6AL translocation and ecosites, as exemplified by identical GMP concentration between the R and S subgroups in the same eco-site, and much higher GMP concentration in Zhengzhou than in Nanjing for the same subgroup.

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Table 3.

X Li et al.

Concentrations of total HMW-GS and GMP in grains of lines with different HMW-GS combination in the two subgroups at different eco-sites

Site and sub-group Nanjing R subgroup

S subgroup

HMW-GS combination

Total HMW-GS concentration (µg mg−1 )

GMP concentration (µg mg−1 )

GS1 (7 + 8, 5 + 10) GS2 (7 + 8, 5 + 12) GS3 (7 + 9, 5 + 10) GS4 (7 + 9, 5 + 12) GS5 (1, 7 + 8, 5 + 10) GS6 (1, 7 + 8, 5 + 12) GS7 (1, 7 + 9, 5 + 10) GS8 (1, 7 + 9, 5 + 12) Mean of R GS1 (7 + 8, 5 + 10) GS2 (7 + 8, 5 + 12) GS3 (7 + 9, 5 + 10) GS4 (7 + 9, 5 + 12) GS5 (1, 7 + 8, 5 + 10) GS6 (1, 7 + 8, 5 + 12) GS7 (1, 7 + 9, 5 + 10) GS8 (1, 7 + 9, 5 + 12) Mean of S

7.12b 7.09a 5.50bc 3.76d 5.55bc 5.90b 4.58c 4.92bc 5.55 ± 1.17 5.98c 4.73d 3.14e 4.83d 7.13b 7.66a 5.99c 4.91d 5.55 ± 1.41 5.55 ± 1.27

49.70a 43.21b 33.47d 22.60e 40.77c 35.56d 40.27c 46.15ab 38.97 ± 8.45 37.58c 36.89c 41.15b 22.63d 40.72b 40.67b 43.58a 37.77c 39.13 ± 2.87 39.05 ± 6.10

GS1 (7 + 8, 5 + 10) GS2 (7 + 8, 5 + 12) GS3 (7 + 9, 5 + 10) GS4 (7 + 9, 5 + 12) GS5 (1, 7 + 8, 5 + 10) GS6 (1, 7 + 8, 5 + 12) GS7 (1, 7 + 9, 5 + 10) GS8 (1, 7 + 9, 5 + 12) Mean of R GS1 (7 + 8, 5 + 10) GS2 (7 + 8, 5 + 12) GS3 (7 + 9, 5 + 10) GS4 (7 + 9, 5 + 12) GS5 (1, 7 + 8, 5 + 10) GS6 (1, 7 + 8, 5 + 12) GS7 (1, 7 + 9, 5 + 10) GS8 (1, 7 + 9, 5 + 12) Mean of S

8.27c 8.27c 7.52d 4.83f 9.77a 8.79b 5.46e 8.34c 7.66 ± 1.64 6.75d 6.26e 4.81f 6.16e 8.83b 10.12a 8.53b 7.4c 7.36 ± 1.67 7.51 ± 1.63

55.89a 41.28d 42.98cd 26.40e 47.19bc 49.01b 42.79cd 45.65c 43.90 ± 8.44 41.61ab 34.69d 39.82b 39.14c 42.35a 41.84a 42.11a 41.82a 40.42 ± 2.58 42.16 ± 6.29

633.60∗∗c 0.02 118.81∗∗ 0.00

23.11∗∗ 6.53∗ 25.01∗∗ 7.89∗∗

Mean in Nanjing Zhengzhou R subgroup

S subgroup

Mean in Zhengzhou F values FE a FP FS FE×P a

FE , FP , FS , FE×P indicates the F value of eco-site, subgroup, subunit and interaction of eco-site by subgroup, respectively. Different superscript letters in the same column mean significant difference at P < 0.05 level. c∗ and ∗∗ refer to significance levels of 0.05 and 0.01, respectively. b

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common wheat usually contains three to five subunits: zero or one encoded by Glu-A1, one or two by Glu-B1 and two by Glu-D1.20 The composition of HMW-GS plays important roles in determining the viscoelastic properties of wheat dough.21 This could be explained by the unique structure of those so-called superior quality HMW-GS, which contain more cysteine residues in the N-terminal domain and strong chain-extending properties


to favour the formation of large glutenin macropolymers22 and gluten.12 In terms of baking quality, it is generally believed that the Glu-1B encoding subunit pair of 7 + 8 is better than 7 + 9,23 and the Glu-1D encoding subunit pair of 5 + 10 is better than 2 + 12,24 and the Glu-1A encoding subunit of 1 or 2∗ is better than null.25 Thus, 1 or 2∗ , 7 + 8 (9), 5 + 10 are recommended as superior quality HMW-GS/pairs for high-quality bread-baking


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HMW-GS concentration (µm g−1)

Eco-site affects wheat grain glutenin accumulation

6

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A

B

8 6

4 4 2 2 0

Nanjing 10

20

30

40

50

60

70

Zhengzhou 10

GMP concentration (µg

20

30

40

50

60

0

70

mg−1)

Figure 4. Relationship between concentrations of GMP and HMW-GS/pairs encoded by genes located at different loci in grains of the RIL population in Nanjing (A) and Zhengzhou (B). The coefficient between the contents of GMP and HMW-GS (pair) 1, 7 + 8, 7 + 9, 5 + 10, 5 + 12 is 0.2016, 0.7147∗∗ , 0.5462∗∗ , 0.6164∗∗ and 0.6586∗∗ , respectively, in Nanjing; and 0.2883, 0.8554∗∗ , 0.7581∗∗ , 0.8012∗∗ and 0.8028∗∗ , respectively, in Zhengzhou. ∗∗ Refers to a significance level of 0.01.

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that the 6 VS·6AL translocation is a potential germplasm for breeding wheat varieties that are resistant to powdery mildew and yellow rust, and there is no risk of reducing/modifying the grain quality. The GMP is a key indicator for wheat quality,9 and mainly consists of HMW-GS and LMW-GS. Our previous experiments14,15 and another study31 have demonstrated that concentrations of GMP and HMW-GS in wheat grain are closely related, and the subunit pair encoded by Glu-1D contributes more to the GMP concentration than do subunit pairs encoded by Glu-1B and Glu1A.14 Here, the correlation coefficient was the highest between the concentration of GMP and HMW-GS pair 5 + 10 and 5 + 12, whilst it was the lowest for subunit 1 in both eco-sites. We further observed that the concentration of GMP and HMW-GS pair 7 + 8 was significantly higher than subunit pair 7 + 9. This further proves that a high concentration due to the presence of the superior quality HMW-GS contributes to a high GMP concentration and hence better wheat grain quality.12 It should be noted that the wheat variety having subunit pair 5 + 10 is reported to contain larger insoluble glutenin (a similar parameter to GMP) than that having subunit pair 2 + 12.24 It is very interesting that the correlation coefficient between concentrations of GMP and HMW-GS pair 5 + 10 was very similar to that of HMW-GS pair 5 + 12 in the present study. Meanwhile, HMW-GS concentrations were not always higher or lower in lines containing subunit pair 5 + 10 than those counterpart lines with pair 5 + 12 as shown in Table 3. This indicated that of the Glu-D1 encoding HMW-GS pairs, y-subunit 10 and 12 could play similar role in affecting HMW-GS concentration, and that the larger insoluble due to the subunit pair 5 + 10 in relation to the pair 2 + 1224 could be more likely ascribed to the variant in the x-subunits (i.e. subunit 5 vs. subunit 2). Of course, more evidence is required to test this conclusion.

CONCLUSION The concentrations of HMW-GS and GMP were largely affected by the eco-sites and the composition of HMW-GS, but not by the 6 VS·6AL translocation. It is suggested that 6 VS·6AL translocation could be potential tool in wheat breeding to improve disease resistance without risk of downgrading grain quality.



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wheat varieties.23,25 It should be also noted that HMW-GS appear to have both qualitative and quantitative effects on grain quality.24 For instance, the transgenic line B73 − 6 − 1, containing 10–15 extra copies of the native HMW glutenin gene (1Dx5), was found to express four-fold more HMW-GS protein and higher ratios of Dx/Dy, HMW-/LMW-GS and glutenin/gliadin, and thereafter resulted in a stronger dough strength.26 In addition, HMW-GS combinations of 1, 7 + 8 (9), 5 + 10 subunits are usually found to possess high HMW-GS concentrations.24 Here, we also proved that the superior quality HMW-GS were associated with higher HMWGS concentration, which could further contribute to the dough quality. Here, RIL lines possessing subunit 1 expressed a higher HMW-GS concentration than those counterpart lines without subunit 1, and the lines having a subunit pair of 7 + 8 contained higher HMW-GS concentration than those having subunit pair of 7 + 9. Therefore, it is suggested that the composition and amount of HMW-GS are closely correlated, and may jointly contribute to dough quality. The baking quality of wheat is not only genetically determined but also affected by its growing environment.27 The accumulation and concentration of HMW-GS in wheat grain is reported to be influenced by nitrogen rate,14 temperature,28 soil moisture15 and other external growth factors.29 In addition, environmental conditions also cause a large variation in HMW-GS concentration and the size distribution of glutenin polymers, and consequently affect the bread-making quality.29 In the present study, we observed that the HMW-GS concentration significantly differed between the two eco-sites, and was much higher in Zhengzhou than in Nanjing. This result indicated that concentration of HMWGS could be site-dependent, which is consistent with earlier findings by Don et al.30 This is also consistent with the fact that the bread-baking quality of wheat from the Huang-Huai-Hai plain is much better than that from the Yangtze River downstream plain in China. The 6 VS·6AL translocation carries genes of Pm21 and Yr26, which are highly resistant to powdery mildew and yellow rust.1 It was very interesting that the HMW-GS concentration in the RIL lines obtained from the cross of Huixianhong with 92R137 (containing 6 VS·6AL translocation) was similar for the R (with 6 VS·6AL translocation) and the S (without 6 VS·6AL translocation) subgroups, indicating that the 6 VS·6AL translocation had no effect on grain HMW-GS concentration, and thus did not affect grain quality, as indicated by Wang et al.8 It is then concluded

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ACKNOWLEDGEMENTS This study was supported by projects of the National Natural Science Foundation of China (31171484, 31028017, 31000686), the Specialized Research Fund for the Doctoral Program of Higher Education (20090097110009), the NCET (06-0493), MATS (nycytx-03) and the Fundamental Research Funds for the Central Universities (KYZ200915). Supporting information Supporting information may be found in the online version of this article.

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J Sci Food Agric 2012; 92: 2188–2194