Ind J Plant Physiol. (January–March 2015) 20(1):97–102 DOI 10.1007/s40502-015-0141-z
SHORT COMMUNICATION
Allelic variations of functional markers for high molecular weight glutenin genes in Indian wheat (Triticum aestivum L.) cultivars and their correlation with bread loaf volume Umesh Goutam • Ratan Tiwari • R. K. Gupta Sarvjeet Kukreja • Ashok Chaudhury
•
Received: 26 May 2014 / Accepted: 1 February 2015 / Published online: 12 February 2015 Ó Indian Society for Plant Physiology 2015
Abstract Glutenins, the polymeric proteins are composed of high molecular weight (HMW) and low molecular weight (LMW) glutenin subunits. High molecular weight glutenin subunit (HMW-GS) plays a key role in determining bread making quality. The HMW-GS encoded by Glu-1 loci (GluA1, GluB1 and GluD1) are highly polymorphic. Thus it is very important to identify the alleles at Glu-1 loci for all three genomes for wheat quality improvement. In the present study 53 Indian wheat genotypes were evaluated for HMW-GS composition using PCRbased DNA markers. In total, two alleles (Ax2*, Ax1 or Axnull) at Glu-A1, four allele/allelic pairs (Bx7, Bx7 ? By8, Bx7 ? By9, Bx17 ? By18) at Glu-B1 and two allelic pairs (Dx2 ? Dy12 and Dx5 ? Dy10) at Glu-D1 were identified using diagnostic DNA markers. Allele or allelic pair Ax2* (62.3 %) at Glu-A1, Bx7 ? By8 (64 %) at Glu-B1 and Dx2 ? Dy12 (58.5 %) at Glu-D1 were present most frequently in wheat genotypes taken for study. Moreover, the allelic combination Ax2*, Bx17 ? By18 and Dx5 ? Dy10 exhibited highest bread loaf volume in comparison to other allelic combinations studied. The set of markers used here thus represents a useful alternative for
U. Goutam R. Tiwari R. K. Gupta Directorate of Wheat Research, Post Box 158, Karnal 132001, India U. Goutam A. Chaudhury Department of Bio and Nanotech, Guru Jambheshwar Univeristy of Science & Technology, Hisar, India U. Goutam (&) S. Kukreja Department of Biotechnology, Lovely Professional University, Phagwara, Punjab, India e-mail:
[email protected]
screening of HMW-glutenin allele (s) for improvement of bread making quality in Indian wheat genotypes. Keywords Triticum aestivum L. HMW glutenin Functional marker Bread loaf volume Among the cereals, wheat is unique because its flour alone has the ability to form dough that exhibits the rheological properties required for the wider diversity of food preparations. In dough, most of the proteins are converted into gluten complex. The gluten proteins consist of glutenin, responsible for elasticity of dough and gliadins, for providing extensibility to dough (Gianibelli et al. 2001; Maucher et al. 2009). Glutenins are composed of high molecular weight (HMW) and low molecular weight (LMW) glutenin subunits (GS) based on their mobility on SDS-PAGE. The HMW-GS are encoded by genes at the Glu-1 loci, present on the long arms of the homeologus group 1 chromosome (Payne and Lawrence 1983). At each Glu-1 locus (Glu A-1, Glu B-1 and Glu-D1), there are two tightly linked HMW-GS genes, which encode two types of subunits, the larger one termed as x-type and the smaller one as y-type (Harberd et al. 1986). Allelic variation existing at each of the HMW glutenin loci demonstrated that dough strength and elasticity is largely influenced by the alleles present at the Glu-D1 locus. The HMW glutenin subunits Dx5 ? Dy10 have been associated with high dough strength and good bread making quality as compared to allelic subunits Dx2 ? Dy12 (Sramkova et al. 2010). In the past, sodium dodecyl sulphate–polyacrylamide gel electrophoresis (SDS-PAGE) has been routinely used to analyze large number of HMW-GS alleles in Indian wheat cultivars (Ram 2003; Singh et al. 2007). However, study by (Gianibelli et al. 2002) has revealed that SDSPAGE analysis of wheat glutenin sometimes results in
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incorrect interpretation of different HMW-GS alleles. The method is time consuming and is not well suited to the high throughput system (Ma et al. 2003). In an effort to overcome these constraints, the availability of PCR-based DNA markers serves as a valuable tool for selection of desired trait with relative ease. The use of PCR-based DNA markers significantly supplements the alternative to SDSPAGE. In last two decades, number of PCR-based DNA markers has been used to differentiate between different HMW-GS alleles (Goutam et al. 2013). The current study thus focuses on the use of identified DNA markers to differentiate between HMW-GS alleles in the bread wheat genotypes grown in different agroclimatic zones of India. Fifty three spring wheat cultivars from five wheat agroclimatic zones of India, viz, North Western Plains Zone (NWPZ), Northern Hills Zone (NHZ), North Eastern Plains Zone (NEPZ), Central Zone (CZ) and Peninsular Zone (PZ) were included in this study. All fifty three wheat cultivars were grown over two crop seasons (2008–2009 and 2009–2010). The experimental material represents the cultivars released during 1965–2011 (Table 1). DNA isolation and PCR analysis was done in Plant Molecular Biology laboratory, Directorate of Wheat Research (DWR), Karnal, India. Genomic DNA was extracted from one-week-old seedlings by SDS method (Dellaporta et al. 1983). The primer sequences, expected product size and annealing temperatures of amplification products are shown in supplementary data. Polymerase chain reaction (PCR) amplifications were carried out in 25 ll volume with STS markers for HMW-GS genes (Ahmad 2000; Ma et al. 2003 and Lei et al. 2006). Bread was baked in Quality and Basic Sciences laboratory, DWR, Karnal, India according to the standard AACC method 26-10 (2000) with slight modifications and loaf volume was measured by the rapeseed displacement method (Dhingra and Jood 2004). A total of 53 bread wheat genotypes were characterized for allelic combinations of HMW-GS allelic composition. Several methods based on biochemical and molecular techniques are available to detect the presence of HMWGS composition (Goutam et al. 2013) in wheat. In the present study, we evaluated the utility of six PCR-based DNA markers for detecting the presence of HMW-GS alleles. All markers were able to amplify the bands that distinguished between wheat genotypes for different HMW glutenin alleles at Glu-1 loci. Using Ax2* allele-specific PCR primers (Ahmad 2000), a band of 1.319 kb was obtained in 33 genotypes, whereas, amplicon was absent in 20 Ax1 or Axnull genotypes. Out of 53 wheat genotypes, 33 genotypes showed the expected product size of 1.319 kb, indicating the presence of Ax2*
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allele, whereas absence of a PCR product in remaining twenty genotypes indicated presence of either Ax1 or Axnull allele (Fig. 1a). Four alleles (Bx7, Bx17, By8 and By9) of Glu-B1 locus were amplified by PCR-based markers developed by Ma et al. (2003) and Lei et al. (2006). PCR analysis using Bx primers (Bx7/Bx17) produced single DNA fragment (669 bp) in genotypes containing the Bx17 allele and two fragments (630 and 766 bp) in genotypes for Bx7 allele/ allelic pair (Fig. 1b). Among the 53 Indian wheat genotypes screened with this marker, thirty four genotypes were found positive for Bx7 allele/allelic pairs (Bx7 ? By8, Bx7 ? By9) and 15 were found positive for Bx17 allele (Table 1). Remaining four genotypes, viz, C 306, HI 1500, HD 2888 and HW 2004 did not show PCR amplicon for Bx7 or Bx17 allele. The genotypes containing By8 allele produced a 527 bp (Fig. 1c) fragment, while non-By8 genotypes were negative for amplicon. Sixteen genotypes showed presence of By8 allele (Table 1). The allele specific primers for identification of different HMW-glutenin allele were used to amplify By9 gene segments. Using By9 AS-PCR primers, a band of 707 bp (genotypes carrying By9 allele) was obtained in 14 genotypes, whereas, amplicon of 662 bp (genotypes carrying non By9 allele) was present in 39 wheat genotypes (Fig. 1d, Table 1). Ma et al. (2003) studied 50 Australian wheat genotypes and reported 39.3 % genotypes with Bx17 allele whereas 36.4 % genotypes were found positive for Bx7 allele. When result of all four AS-PCR (Table 1) primers was collectively analyzed it was found that 64 % genotypes showed presence of Bx7allele/allelic pairs (Bx7, Bx7 ? By8, Bx7 ? By9) while remaining 28 % genotypes showed presence of Bx17 allele (Table 1). This data indicates that Bx7 allelic pair is more prominent in Indian wheat genotypes as compared to Bx17 allele. A multiplex PCR assay for fifty three wheat genotypes was carried out which permitted the discrimination of the major HMW glutenins in a single PCR reaction and agarose gel assay. Primer combinations (Dx5 and Dy10/12) were used to establish a simple multiplex PCR assay to distinguish both alleles of HMW glutenin, 1Dx and 1Dy, respectively (Fig. 1e). In the present study, 22 genotypes amplified 450 and 576 bp fragments, indicating the presence of Dx5 and Dy10 alleles, respectively. Remaining 31 genotypes amplified 612 bp fragment associated with Dy12 allele (Table 1), whereas absence of Dx5 allele indicating presence of Dx2 allele. The overall frequencies of allelic pairs Dx5 ? Dy10 and Dx2 ? Dy12 were 41.5 and 58.5 %, respectively. It shows that Dx2 ? Dy12 allelic pair is predominantly present in Indian wheat genotypes. Table 1 shows the range of variation of bread loaf volume (BLV) of 53 bread wheat genotypes studied. The BLV varied from 498 ml (HI 1500) to 606 ml (HI 977) with an
HS 490
HS 507
Sonalika
32
33
34
HI 1563
22
VL 892
HD 2967
21
HS 420
MACS 6145 HD 2888
19 20
31
K 0307
18
30
NW 1014
17
HS 295
HUW 234
16
VL 829
K 8027
15
29
NW 2036
14
28
DBW 14
13
VL 829
HD 2733
12
28
K 9107
11
HS 277
C 306
10
27
HI 1544
9
VL 616
HW 2004
8
VL 804
HI 1500
7
26
Raj 4037
6
25
GW 173
5
HS 240
MP 4010 DL 788-2
3 4
VL 738
GW 322
2
23
Lok 1
1
24
Cultivar
S.no.
II54-388/AN/3/YT54/N10B/LR 64
KAUZ/MYNA/VUL/BUC/FLK/4/MILAN
HS 364/HPW 114//HS 240/HS 346
WH 542/PBW 226
LAJ 3302//CMH 73A-497/3*CNO 79
(OT-AZ/IA 555-ALD’’S‘‘/ALD’’S‘‘-NATN) PJN’’S‘‘-PEL ’’S‘‘.
BWSN/CPAN2099
BWSN/CPAN2099
KVZ/CGN
SONALIKA/CPAN 1507
CPAN 3018/CPAN 3004//PBW 65
NS12.07/LIRA’’S’’//VEE ,,S’’
AU/KAL-BB//WOP’’S‘‘/PAVON’’S‘‘
MACS 2496*2/MC 10
ALD/COC//URES/HD2160 M/HD2278
C306 ? LR28 C 306/T. sphaerococcum//HW 2004
K 8321/UP 2003
HAHN ,,S‘‘
HUW 12* 2/CPAN 1666//HUW 12
NP875/4/N10B/Y53//Y50/3/KT54B/5/2*K852
BOW/CROW/BUC/PVN
RAJ 3765/PBW 343
ATTILA/3/TUI/CARC//CHEN/CHTO/4/ATTILA
K 8101/K 68
RGN/CSK3//2*C591/3/C217/N14//C281
HINDI 62/BOBWHITE/CPAN 2099
C 306 *7//TR 380-14 #7/3 AG14
HW2002*2//STREMPALLI/PNC 5
DL 788-2/RAJ 3717
TW275/7/6/1/LOK-1
ANGOSTURA 88 K7537/HD2160/HD2278//L24/K4.14
GW 173/GW 196
S308/S331
Pedigree
NWPZ
NHZ
NHZ
NHZ
NHZ
NHZ
NHZ
NHZ
NHZ
NHZ
NHZ
NHZ
NHZ
NEPZ
NEPZ
NEPZ NEPZ
NEPZ
NEPZ
NEPZ
NEPZ
NEPZ
NEPZ
NEPZ
NEPZ
NEPZ
CZ
CZ
CZ
CZ
CZ
CZ CZ
CZ
CZ
Zone
1965
2010
2007
2007
2002
1992
2002
2002
1992
1986
2002
1996
1989
2010
2011
2002 2005
2006
1997
1985
1984
2002
2002
2001
1995
1965
2007
1995
2002
2003
1994
2002 1996
2002
1981
Year of release
?
–
?
?
?
?
–
–
–
?
?
–
–
?
?
? –
?
?
?
?
–
?
?
?
–
–
–
?
?
?
? –
?
?
Ax2*
?
?
?
?
?
–
?
?
?
?
?
?
?
?
–
– –
–
?
?
–
?
?
?
–
–
?
–
–
–
?
– –
?
–
BX7
–
–
–
–
–
?
–
–
–
–
–
–
–
–
?
? –
?
–
–
?
–
–
–
?
–
–
–
–
?
–
? ?
–
?
BX17
–
?
?
?
?
–
–
–
–
–
–
?
–
?
–
– –
–
?
?
–
?
?
–
–
–
?
–
–
–
?
– –
?
–
BY8
?
–
–
–
–
–
?
?
?
?
?
–
?
–
–
– –
–
–
–
–
–
–
?
–
–
–
–
–
–
–
– –
–
–
BY9
?
–
?
?
?
?
?
?
–
?
?
–
–
?
–
– ?
?
?
?
?
–
?
–
–
?
?
?
?
?
?
? ?
?
–
Dx2 ? Dy12
Table 1 Allelic variation of HMW glutenin genes at Glu-A1, Glu-B1, Glu-D1 loci and mean bread loaf volume (BLV) of 53 Indian wheat genotypes
–
?
–
–
–
–
–
–
?
–
–
?
?
–
?
? –
–
–
–
–
?
–
?
?
–
–
–
–
–
–
– –
–
?
Dx5 ? Dy10
531
557
517
532
534.5
515.3
520
520
521.8
512
523.3
558
549.3
555
568
502.8 521.5
603
538
547
538.8
598
543.8
576.3
599.5
503.3
538
519.5
498
573
567.7
560 564
535.3
573.5
BLV (ml)
Ind J Plant Physiol. (January–March 2015) 20(1):97–102 99
123
123
UP 2425
PBW 175
PBW 396
DBW 17
PBW 502 PBW 550
PBW 590
HD 2329
NI 5439
Raj 4083
HD 2189
40
41
42
43
44 45
46
47
48
49
50
HD 2932
Raj 3765
39
53
PBW 373
38
NIAW 34
PBW 343
37
HI 977
HD 2687
36
51
WH 542
35
52
Cultivar
S.no.
Table 1 continued
KAUZ/STAR//HD 2643
GLL/AUST 61.157/CNO/NO 66/3/Y50E/3/KAL
CNO 79/PRL ‘‘S’’
HD 1963/HD 1931
PBW 343/UP 2442//WR 258/UP 2425
REMP 80/3* NP 710
HD 1962/E 4870/3/K 65/5/HD1553/4/UP262
WH 594/RAJ3814//W 485
W 485/PBW 343//RAJ 1482 WH 594/RAJ 3858//W 485
CMH79A.95/3*CNO 79//RAJ3777
CNO67/MFD//MON’’S’’/3/SERI
HD 2160/WG 1025
HD 2320/UP 2263
HD 2402/VL639
ND/VG9144//KAL/BB/3/YCO’’S‘‘/4/VEE#5 ’’S‘‘
ND/VG9144//KAL/BB/3/YCO’’S‘‘/4/VEE#S ’’S‘‘
CPAN2009/HD 2329
JUP/BJY’’S’’//URES
Pedigree
PZ
PZ
PZ
PZ
PZ
PZ
NWPZ
NWPZ
NWPZ NWPZ
NWPZ
NWPZ
NWPZ
NWPZ
NWPZ
NWPZ
NWPZ
NWPZ
NWPZ
Zone
2007
1985
1995
1979
2006
1973
1982
2008
2003 2007
2006
1999
1989
1999
1995
1996
1995
1999
1992
Year of release
?
?
–
?
–
–
?
?
– ?
?
–
?
–
–
–
–
?
?
Ax2*
–
–
?
–
?
–
?
?
? ?
?
?
?
–
?
?
?
?
?
BX7
?
?
–
?
–
?
–
–
– –
–
–
–
?
–
–
–
–
–
BX17
–
–
–
–
?
–
–
–
– –
–
–
?
–
?
–
–
–
–
BY8
–
–
?
–
–
–
?
?
– ?
–
?
–
–
–
–
–
?
?
BY9
?
–
–
?
–
?
?
–
– –
–
–
?
–
?
–
–
?
–
Dx2 ? Dy12
–
?
?
–
?
–
–
?
? ?
?
?
–
?
–
?
?
–
?
Dx5 ? Dy10
585
606.5
535.3
577.5
590.3
592
500
562
550.3 544
564.5
549.3
512.8
560
542
557
544.5
526.5
542.8
BLV (ml)
100 Ind J Plant Physiol. (January–March 2015) 20(1):97–102
Ind J Plant Physiol. (January–March 2015) 20(1):97–102
Fig. 1 a PCR amplification of HMW glutenin Ax2* gene with allelespecific PCR marker in bread wheat genotypes. 1 DBW 14, 2 DBW 17, 3 GW 173, 4 GW 322, 5 HD 2189, 6 HD 2733, 7 VL 616, 8 HS 490, 9 HI 977, 10 HS 420, 11 HW 2004, 12 K 0307, 13 K 8027, 14 1-Kb DNA ladder. b PCR amplification of HMW glutenin alleles Bx7/Bx17 in bread wheat genotypes with allele specific-PCR marker related to bread quality. 1 DL 788-2, 2 GW 173, 3 HD 2189, 4 HD 2687, 5 HS 295, 6 HS 277, 7 C 306, 8 HS 420, 9 K 0307, 10 NIAW 34, 11 NI 5439, 12 PBW 175, 13 100–1,200 bp DNA ladder. c PCR amplification of HMW glutenin By8 gene with allele-specific PCR marker in bread wheat genotypes. 1 HI 977, 2 HD 2329, 3 C 306, 4 DBW 17, 5 DBW 14, 6 DL 788-2, 7 HD 2189, 8 GW 173, 9 HD 2687, 10 HD 2733, 11 HI 1500, 12 HS 240, 13 HS 277, 14 GW 322, 15 HS 295, 16 HW 2004, 17 K 0307, 18 K 8027, 19 HS 420, 20 100 bp DNA ladder. d PCR amplification of HMW glutenin By9 gene with allelespecific PCR marker in bread wheat genotypes. 1 HI 977, 2 HD 2329, 3 HS 295, 4 K 0307, 5 HD 2733, 6 K 8027, 7 NIAW 914, 8 1.2 Kb DNA ladder. e Multiplex PCR for amplification of HMW glutenin 1DX5 and 1Dy10/1Dy12 alleles with 450 and 576/612 bp DNA fragments, respectively from bread wheat genotypes. 1 100 bp DNA ladder, 2 PBW 343, 3 HI 977, 4 PBW 373, 5 NIAW 34, 6 HW 2004, 7 HI 1500, 8 MACS 6145, 9 RAJ 3765
average value of 547.8 ml. Higher BLV (more than 575 ml) was found only in nine genotypes (K0 307, HD 2189, HD 2932, NI 5439, NW 2036, K 9107, RAJ 4083, HD 2733 and HI 977), 14 genotypes were found to possess
101
moderate BLV (550–575 ml), whereas, 30 genotypes showed poor (525–550 ml) and inferior values of BLV (\525 ml). Out of 33 genotypes (carrying Ax2* allele), only six genotypes viz., HI 977, K 9107, K 0307, HD 2189, HD 2932 and HD 2733 showed higher BLV (Table 1). Inspite of the presence of Ax2* allele in remaining twenty seven genotypes, BLV was moderate to low. It might be because of the fact that out of 27 genotypes, 20 genotypes were also carrying the Dx2 and Dy12 alleles responsible for poor bread making quality (Payne and Lawrence 1983) whereas, remaining seven genotypes showed moderate value of BLV with presence of favorable alleles Dx5 and Dy10. At Glu-B1 locus, 34 genotypes carrying Bx7 ? By8 and Bx7 ? By9 allele/allelic pairs, 31 genotypes showed moderate to lower values of BLV (Table 1). Out of these 31 genotypes, 18 genotypes showed poor BLV and 13 genotypes showed moderate value with presence of allelic pair Dx2/Dy12 and Dx5/Dx10, respectively (Payne and Lawrence 1983). Out of 15 genotypes carrying Bx17 ? By18 allelic pair, only three genotypes, viz, HS 295, K 8027 and MACS 6145 was found with poor BLV (Table 1). Two genotypes, viz, HS 295 and K 8027 were also carrying poor bread making alleles Dx2 and Dy12. Lawrence et al. (1987) could not find any difference in dough quality between Bx7 ? By8 and Bx17 ? By18 subunits but they reported that Bx17 ? By18 provided greater loaf volume and was as effective as Dx5 ? Dy10 for improving dough and baking quality (Lawrence et al. 1987). At Glu-D1 locus, out of 22 genotypes carrying Dx5 ? Dy10 allelic pair, only two genotypes, viz, MACS 6145 and HS 277 showed inferior value of BLV, whereas six genotypes, viz., HS 240, WH 542, PBW 343, PBW 396, PBW 550 and NIAW 34 were found with reasonably poor values of BLV. Out of these eight genotypes having poor BLV, seven genotypes were not carrying favorable alleles at Glu-B1 loci (Bx17 ? By18) and one genotype MACS 6145 exceptionally showed poor BLV, inspite of having all the favorable alleles. Out of 31 genotypes (carrying Dx2 ? Dy12 allelic pair), only four genotypes namely, K 0307, NI 5439, HD 2932 and HD 2189 showed higher values of BLV (Table 1). It might be because of the fact that these genotypes also have other favorable allelic pair Bx17 ? By18 at Glu-B1 locus, which contributed positively as Dx5 ? Dy10 allelic pairs for bread quality (Lawrence et al. 1988). The effect of interaction of HMW-glutenin allelic combination on BLV was also analyzed. The relationship between different combination of HMW-glutenin subunits and bread loaf volume is shown in Fig. 2. The highest mean value was observed for HMW-GS combination Ax2*, Bx17 ? By18 and Dx5 ? Dy10, whereas Ax2*,
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Bread loaf volume (ml)
600 580 560 540 520 500 17+18, 5+10 17+18, 2+12
7+9, 5+10
7+8, 2+12
7+9, 2+12
HMW glutenin allelic combination
Fig. 2 Relationship between different combination of HMW-glutenin subunits and bread loaf volume of 53 bread wheat genotypes
Bx7 ? By9 and Dx2 ? Dy12 subunits combination showed lowest mean value. Out of 53 genotypes, only five genotypes, viz, HI 977, K 9107, HD 2967, LOK 1 and MACS 6145 showed favorable allelic combination (Ax2*, Bx17 ? By18 and Dx5 ? Dy10) in Indian wheat genotypes. This indicates that HMW-GS Dx5 ? Dy10 are superior to Dx2 ? Dy12 for good bread making properties (Payne and Lawrence 1983; Gupta and MacRitchie 1991; Luo et al. 2001).
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Goutam, U., Kukreja, S., Tiwari, R., Chaudhury, A., Gupta, R. K., Dholakia, B. B., & Yadav, R. (2013). Biotechnological approaches for grain quality improvement in wheat: Present status and future possibilities. Australian Journal of Crop Science, 7, 469–483. Gupta, R. B., & MacRitchie, F. (1991). A rapid one-step onedimensional SDSPAGE procedure for analysis of subunit composition of glutenin in wheat. Journal of Cereal Science, 14, 105–109. Harberd, N. P., Bartels, D., & Thompson, R. D. (1986). DNA restriction fragment variation in the gene family encoding highmolecular-weight (HMW) glutenin subunits of wheat. Biochemical Genetics, 24, 579–596. Lawrence, G. J., MacRitchie, F., & Wrigley, C. W. (1988). Dough and baking quality of wheat lines deficient in glutenin subunits controlled by the Glu-Al, Gut-B1 and Glu-D1loci. Journal of Cereal Science, 7, 109–112. Lawrence, G. J., Moss, H. J., Shepherd, K. W., & Wrigley, C. W. (1987). Dough quality of biotypes of eleven Australian wheat cultivars that differ in high-molecular-weight glutenin subunit composition. Journal of Cereal Science, 6, 99–101. Lei, Z. S., Gale, K. R., He, Z. H., Gianibelli, C., Larroque, O., Xia, X. C., et al. (2006). Y-type gene specific markers for enhanced discrimination of high-molecular weight glutenin alleles at the Glu-B1 locus in hexaploid wheat. Journal of Cereal Science, 43, 94–101. Luo, C., Griffin, W. B., Branlard, G., & McNeil, D. L. (2001). Comparison of low and high molecular weight wheat glutenin allele effects on flour quality. Theoretical Applied Genetics, 102, 1088–1098. Ma, W., Zhang, W., & Gale, K. R. (2003). Multiplex-PCR typing of high molecular weight glutenin alleles in wheat. Euphytica, 134, 51–60. Maucher, T., Figueroa, J. D. C., Reule, W., & Pena, R. J. (2009). Influence of low molecular weight glutenins on viscoelastic properties of intact wheat kernels and their relation to functional properties of wheat dough. Cereal Chemistry Journal, 86, 372–375. Payne, P. I., & Lawrence, G. J. (1983). Catalogue of alleles for the complex gene loci, Glu-A1, Glu-B1, and Glu-D1 which code for high-molecular weight subunits of glutenin in hexaploid wheat. Cereal Research Communications, 11, 29–35. Ram, S. (2003). High molecular weight glutenin subunit composition of Indian wheats and their relationships with dough strength. Journal of Plant Biochemistry and Biotechnology, 12, 151–155. Singh, A. M., Devaeshwar, J. J., Ahlawat, A. K., & Singh, B. B. (2007). Identification of novel variants of high molecular weight glutenin subunits in Indian bread wheat landraces. Cereal Research Communications, 35, 99–108. Sramkova, Z., Gregova, Z., Slikova, S., & Studike, E. (2010). Wheat varieties released in Slovakia and their bread-making quality. Cereal Research Communications, 38, 386–394.