Barley can be classified as two-row and six-row by vyo46383


									           Comparisons of Two-Row and Six-Row Barley for Chemical Composition
                              Using Doubled-Haploid Lines
            Judith Fregeau-Reid, Thin-Meiw Choo,* Keh-Ming Ho, Richard A. Martin, and Takeo Konishi
                           ABSTRACT                                                protein content than six-row genotypes (Day and Dick-
   Comparative studies on chemical composition between two-row                     son, 1957; Pomeranz et al., 1973; Hockett and Stand-
and six-row barley (Hordeum vulgare L.) and between purple and                     ridge, 1976; Takahashi et al., 1976; McGuire and Hock-
yellow barley are very limited. Therefore, a study was conducted to                ett, 1983; Marquez-Cedillo et al., 2000). Comparing 32
compare two-row and six-row barley and to compare purple and                       two-row (vrs1.b) cultivars with 43 six-row cultivars, Nar-
yellow barley for five chemical traits. In addition, the effects of four           asimhalu et al. (1995) found that -glucan content, on
other marker loci—srh (short rachilla hair), Raw1 (rough awn), Est1                average, was lower for two-row than six-row barleys;
(esterase 1), and Est5 (esterase 5)—on the five traits were also studied.
                                                                                   and Kong et al. (1995) reported that two-row cultivars
One hundred ninety doubled-haploid (DH) lines were derived from
a ‘Leger’/‘CI9831’ cross by the bulbosum method. The DH lines and
                                                                                   on average contained more starch and less fiber than
the two parents were evaluated for protein, starch, -glucan, neutral-              six-row cultivars. The findings of these two studies, how-
detergent fiber (NDF), and acid-detergent fiber (ADF) content at                   ever, were confounded by different genetic backgrounds
two locations in Eastern Canada in 1993. Results showed that two-                  between the two types of barley. To our knowledge, no
row (vrs1.t ) lines contained 14 to 20% more protein, 4% less starch,              one has studied the effect of the deficient allele vrs1.t
and 6 to 7% more -glucan than six-row lines; while purple lemma                    on protein, -glucan, starch, and fiber content in barley.
(Pre2 ) lines contained 2 to 4% less NDF and 0 to 3% less ADF than                    Barley can also be classified as black, purple, and yel-
yellow lemma lines. Differences in grain protein, starch, and -glucan              low (white) on the basis of lemma and pericarp color.
content were associated with the Pre2 locus, but they were shown to                Syrian farmers have firmly believed that a black-seeded
be caused by linkage between the Pre2 and vrs1 loci. Alleles at the                landrace “provides better feed for sheep than the white-
srh, Raw1, Est1, and Est5 loci had very little effect on the five traits.
                                                                                   seeded landrace” (Ceccarelli and Grando, 1999). The two
Protein content was not correlated with -glucan content for either
two-row or six-row lines. Protein and -glucan content, however,
                                                                                   types of landrace, however, showed no difference in pro-
were correlated with NDF and ADF content for the two-row lines.                    tein and lysine content (Ceccarelli and Grando, 1999).
Additive additive epistasis was detected for starch and NDF content.               On the other hand, black barley was reported to contain
The results of this study suggested that selection for high protein or             more protein, crude fiber, riboflavin, lysine, and calcium
low -glucan is possible in two-row/six-row crosses.                                than white barley (Li et al. 1999). To our knowledge, no
                                                                                   comparative studies on chemical composition between
                                                                                   purple and yellow barley have been published in the lit-
B    arley can be classified as two-row and six-row
      types on the basis of spike morphology. The differ-
ence in spike type is controlled primarily by a single
                                                                                      Doubled-haploid (DH) lines are ideal for genetic anal-
                                                                                   ysis because they represent a random set of completely
gene vrs1 with the two-row type being dominant. This                               homozygous lines. Therefore, DH lines derived from a
vrs1 gene has remarkable effects on many other charac-                             two-row/six-row cross were used in this investigation.
teristics. Recently, Jui et al. (1997) reported that six-                          Our objectives were to determine the effects of the vrs1.t
row lines outyielded deficient two-row (vrs1.t) lines by                           and Pre2 alleles on five chemical traits (protein, starch,
20 to 27% in Eastern Canada but that two-row lines                                   -glucan, NDF, and ADF contents), investigate the po-
had higher test and seed weight and better resistance                              tential for obtaining transgressive lines for the five chem-
to lodging. Two-row (vrs1.t) lines also produced larger                            ical traits, and determine if additive additive epistasis
and more circular seeds than did six-row lines (Fregeau-                           (i.e., homozygote       homozygote interaction) and ge-
Reid et al., 1996). Since two-row barley produces larger                           netic correlations are present for the five chemical traits
seeds with higher test weight and seed weight than six-                            in a two-row/six-row barley cross. Data for four other
row barley, it is very likely that two-row barley produces                         marker loci—srh (short rachilla hair), Raw1 (rough awn),
better quality feed than six-row barley. In fact, Bouard                           Est1 (esterase 1), and Est5 (esterase 5)—were also avail-
et al. (1980) found that two-row (vrs1.b) barley had                               able, and thus they were used to study the effects of
4.6% better feed conversion than six-row barley in a                               these four other markers on the five chemical traits.
swine feeding trial. Comparative studies on chemical
composition between two-row and six-row barley are                                           MATERIALS AND METHODS
very limited. Two-row (vrs1.b) genotypes had higher
                                                                                                          Plant Materials
                                                                                     One hundred ninety DH lines were derived from the F1
J. Fregeau-Reid, T.M. Choo, and K.M. Ho, Eastern Cereal and Oil-
seed Research Centre, Agriculture and Agri-Food Canada, Ottawa,                    hybrids of a Leger/CI9831 cross of barley by the bulbosum
ON, Canada K1A 0C6; R.A. Martin, Crops and Livestock Research                      method (Choo et al., 1992). Leger is a six-row cultivar well
Centre, Agriculture and Agri-Food Canada, P.O. Box 1210, Char-                     adapted to Eastern Canada while CI9831 is a two-row intro-
lottetown, PEI, Canada C1A 7M8; T. Konishi, 294 Okada, Mabi-                       duction resistant to net blotch, caused by Pyrenophora teres
cho, Kibi-gun, Okayama 710-1311, Japan. Received 30 Oct. 2000.
*Corresponding author (
                                                                                   Abbreviations: DH, doubled haploid; QTL, quantitative trait loci;
Published in Crop Sci. 41:1737–1743 (2001).                                        NDF, neural-detergent fiber; ADF, acid-detergent fiber.

1738                                          CROP SCIENCE, VOL. 41, NOVEMBER–DECEMBER 2001

Table 1. Classification of the 190 doubled-haploid (DH) lines                                    Statistical Analysis
  derived from a Leger/CI9831 cross of barley into 12 marker
  classes on the basis of six marker loci.                                      Data from each location were analyzed separately. The
                                                                            average of the duplicate samples was used for each experimen-
                                            No. of DH lines
                                                                            tal unit. The mean of the 190 DH lines was compared with
Locus       Chromosome          Leger genotype†        CI9831 genotype      the mid-parent by an F-test to detect whether or not additive
vrs1             2H                    96                      94           additive epistasis was present for each trait (Choo et al., 1986).
Pre2             2H                    93                      97           In theory, the two means should be the same in the absence of
Raw1             5H                    94                      96           additive additive epistasis (Choo et al., 1986). The frequency
Srh              5H                    91                      99           distribution of the 190 DH lines was tested for normality by
Est1‡            3H                    71                      62
Est5‡            7H                    83                      50           the W-test (Shapiro and Wilk, 1965) to determine if additive
                                                                            additive epistasis was present for the chemical traits. The fre-
† Leger carries the vrs1.a (six-row), pre2 (yellow lemma), srh (short ra-   quency distribution should be normal in the absence of
  chilla hair), raw1 (smooth awn), Est1.Ca, and Est5.Te alleles. CI9831
  carries the vrs1.t (two-row), Pre2 (purple lemma), Srh (long rachilla     additive additive epistasis (Choo and Reinbergs, 1982). DH
  hair), Raw1 (rough awn), Est1.Pr, and Est5.Ri alleles.                    lines were compared with each parent by the t-test. The pooled
‡ The allelic identity of the other 57 lines was not determined.            mean square among the five entries for each parent was used
                                                                            as error mean square for starch and -glucan content.
(Died.) Dreschsl. (Ho et al., 1996). Leger carries the vrs1.a,                  Phenotypic correlation was used to detect genetic linkage
pre2, srh, raw1, Est1.Ca, and Est5.Te alleles. In contrast,                 and/or pleiotropy. Assuming Traits X and Y with heritabilities
CI9831 (which was obtained from the Plant Gene Resources                    h2 and h2 , respectively, Falconer (1960) showed that pheno-
                                                                              X        Y

of Canada) carries the vrs1.t, Pre2, Srh, Raw1, Est1.Pr, and                typic correlation (rp) between X and Y is a linear combination
Est5.Ri alleles. The 190 DH lines can be classified into 12                 of genetic correlation (rA) and environmental correlation (r E ).
marker classes according to the six marker loci (Table 1).                  Mathematically, rp       hX h Y rA   eX eY r E where e2
                                                                                                                                  X   1    h2X

The Est5 gene segregated distortedly, but all the other genes               and e2 Y     1   h2 (see Falconer 1960, Eq. [19.1]). When an

segregated at random (Choo et al., 1992). The vrs1 and Pre2                 identical set of DH lines is grown at locations as diverse as
loci were linked in this set of DH lines with a recombination               Charlottetown (C) and Ottawa (O), then the environmental
value of 0.11, while the Srh and Raw1 loci were linked with                 correlation for X and Y becomes negligible or zero (i.e., r E
a recombination value of 0.30 (Jui et al., 1997). The two parents           0). Consequently, the resulting phenotypic correlation is due
and the 190 DH lines were seeded at Charlottetown (Prince                   solely to the genetic correlation. By this method, two pheno-
Edward Island) and Ottawa (Ontario) in 1993, in a randomized                typic correlation coefficients between X and Y were obtained,
complete block design with four replicates at each location                 one was between X at C and Y at O and the other was between
(see Jui et al., 1997 for details). The two parents were each               X at O and Y at C. The two correlation coefficients varied
represented five times in each replicate. Standard cultural prac-           somewhat dependent upon the heritability values. Correlation
tices were followed at each location.                                       analysis was conducted for the 94 two-row lines and for the
                                                                            96 six-row lines to detect linkages of loci (except the vrs1
                        Chemical Analysis                                   locus) affecting two chemical traits.
                                                                                The effects of the six marker loci on chemical traits were
   Seed samples from two replicates were analyzed for protein               studied by comparing the two marker classes for each of the
and NDF and ADF content, while seed samples from one                        six marker loci. The between classes mean square was tested
of the two replicates were analyzed for starch and -glucan                  against the within classes mean square by the F-test (Choo,
content. Each sample was analyzed in duplicate. Seed samples                1983). If means of the two marker classes of DH lines were
used for chemical analyses were ground to pass through a                    significantly different, then this would suggest the presence
0.5-mm screen on a Udy-mill. Moisture content was measured                  of either linkage between the marker locus and quantitative
following the air-oven method, AACC method 44-15A                           trait loci (QTL), or a pleiotropic effect of the marker gene. The
(AACC, 2000). All results were expressed on a dry weight                    variance component between classes and variance component
basis. Protein content (N 6.25) was determined with a Teca-                 within classes were estimated by equating the expected mean
tor Kjeltec Auto Analyzer 10301 (Tecator AB, Hoganas, Swe-
                                                 ¨   ¨                      squares to their observed mean squares. The effect of the
den) with the macro-kjeldahl unit attachment. Starch and                    marker genes were further studied by determining the percent-
  -glucan content were determined enzymatically following the               age of the variance component between classes over the total
barley grains procedures of the commercial kits from Mega-                  variance (i.e., sum of between classes and within classes vari-
zyme (Megazyme International Ireland Ltd., Bray, County                     ance components). The effects of the two chromosome seg-
Wicklow, Ireland); the starch protocol was AACC method                      ments, one flanked by the vrs1 and Pre2 loci and the other
76-13 (AACC, 2000) and the -glucan protocol was AACC                        by the Srh and Raw1 loci, on the five chemical traits were
method 32-23 (AACC, 2000). NDF and ADF content were                         studied by comparing the means and variances of the four
measured following slightly modified protocols (incubation                  marker classes in two by two factorials (Choo, 1983). Bartlett’s
time for NDF 80 min. and for ADF 70 min.) developed                         and the F-tests were used to determine if the four variances
by ANKOM Technology Corp. with the ANKOM 200 Fiber                          were homogeneous (Snedecor and Cochran, 1967).
Analyzer (ANKOM Technology Corp., Fairport, NY). Before
fiber determinations, starch was removed by overnight incuba-
tion at 40 C in a phosphate buffered solution with alpha-                              RESULTS AND DISCUSSION
amylase at pH       7.0. All chemicals and the alpha-amylase
enzyme were ANKOM brand purchased through Anachemia                                        Potential of Lines
Canada Inc.                                                                    At Charlottetown, CI9831 contained 41% higher pro-
                                                                            tein than Leger and it contained a similar amount of
     Mention of product and equipment names is for identification
                                                                            starch, -glucan, NDF, and ADF as Leger (Table 2).
purposes only and does not imply a warranty or endorsement to the           At Ottawa, CI9831 contained 38% higher protein than
exclusion of other products that may be similar.                            Leger but Leger contained 7% higher starch, 12%
                                         FREGEAU-REID ET AL.: CHEMICAL COMPOSITION OF BARLEY                                            1739

Table 2. Means and tests for normality for 190 doubled-haploid               Table 3. Correlation coefficients between chemical traits within
  (DH) lines derived from a Leger/CI9831 cross of barley grown                 two-row and six-row doubled-haploid lines derived from a
  at two Eastern Canada locations.                                             Leger/CI9831 cross of barley grown at two Eastern Canada
Parent/DH         Protein     Starch      -glucan       NDF        ADF
                                          g kg
Charlottetown                                                                Charlottetown       Protein    Starch   -glucan   NDF      ADF
  Leger           128a†        526a       48a          149a        66a
  CI9831          180b         520a       41a          151a        65a       94 Two-row lines
  DH              161         (501)‡      45          (160)       (72)         Protein                       0.23*   0.03      0.23*    0.30**
  W statistic       0.98         0.99      0.99          0.95**     0.96**     Starch             0.07               0.22*     0.00     0.04
Ottawa                                                                           -glucan          0.17       0.03              0.43**   0.35**
  Leger            112a        570a       56a          149a       70a          NDF                0.22*      0.23*   0.32**             0.43**
  CI9831           155b        532b       50b          150a       63b          ADF                0.27**     0.23*   0.25*     0.46**
  DH              (145)       (505)      (45)         (159)       70         96 Six-row lines
  W statistic§       0.96**      0.99      0.96**        0.97      0.97        Protein                       0.17    0.13      0.07     0.16
                                                                               Starch             0.30**             0.12      0.02     0.16
** Significant deviation from normality at the 0.01 level.                       -glucan          0.07       0.13              0.17     0.17
† Means for the same trait followed by different letters were different at     NDF                0.04       0.21*   0.41**             0.52**
  the 0.05 level.                                                              ADF                0.12       0.16    0.38**    0.36**
‡ Means in parentheses were different from their respective mean values
  of parents at the 0.05 level.                                              * Indicates significance at P 0.05.
§ Shapiro-Wilk W statistic.                                                  ** Indicates significance at P 0.01.

higher -glucan, and 11% higher ADF than CI9831.                              lower than the mid-parent for starch content at both
The two parents were not different in NDF content at                         locations and for -glucan content at Ottawa (Table 2).
Ottawa. Four lines contained higher protein than                             On the other hand, the mean of the DH lines was higher
CI9831 at Charlottetown. Two of the four lines were                          than the mid-parent for protein content at Ottawa, NDF
two-row (191 and 198 g protein kg 1 ) and the other two                      content at both locations, and ADF content at Char-
were six-row (193 and 195 g protein kg 1 ). Twenty-four                      lottetown. The frequency distributions of the 190 DH
lines contained higher protein than CI9831 at Ottawa.                        lines for protein content appeared to be bimodal while
Twenty-three of them were two-row (164 to 181 g pro-                         those for the other four traits appeared to be unimodal
tein kg 1 ) and only one was six-row (164 g protein kg 1 ).                  at both locations (data not shown). The frequency distri-
One two-row line contained the highest amount of pro-                        bution was nonnormal for protein and -glucan content
tein at both locations. No lines contained less protein                      at Ottawa and for NDF and ADF content at Charlotte-
than Leger at either location. None of the 190 lines                         town (Table 2). Both the normality method (which was
contained more starch than Leger and less ADF than                           based on the DH lines only) and the mean comparison
CI9831. One six-row line contained the lowest -glucan                        method (which was based on both DH lines and their
at Charlottetown (23 g kg 1 ) and the second lowest at                       parents) detected additive additive epistasis for NDF
Ottawa (29 g kg 1 ). On the other hand, one two-row                          and ADF content at Charlottetown and for protein and
line contained more -glucan than Leger at Char-                                -glucan content at Ottawa. Of the two methods, the
lottetown (65 g kg 1 ). One two-row line contained less                      mean comparison method should be more accurate be-
NDF than Leger at either location. Many of these DH                          cause theoretically it has a smaller standard error (Ken-
lines contained more NDF than CI9831 with the major-                         dall and Stuart, 1967). The results indicated that addi-
ity of them being six-row lines (36 vs. 31 at Char-                          tive additive epistasis was present at least for starch
lottetown and 60 vs. 33 at Ottawa). In this study, CI9831                    and NDF content, and perhaps for protein, -glucan,
contained 38 to 41% higher protein than Leger. Still,                        and ADF content also. To our knowledge, additive
DH lines with higher protein content than CI9831 were                        additive epistasis has not been reported for these chemi-
found in this cross, indicating that the low-protein parent                  cal traits previously. If additive    additive epistasis is
Leger also carried high protein genes. Previously, Mar-                      indeed important for these chemical traits, then selec-
quez-Cedillo et al. (2000) reported that the largest QTL                     tion for these chemical traits should not be too severe in
for protein content was coincident with the vrs1 locus,                      early stages of a breeding program for allowing desirable
but there were also four minor QTL on four different                         epistatic combinations to come together.
chromosomes (2H, 4H, 5H, and 7H) in the ‘Harrington’
(two-row)/‘Morex’ (six-row) cross. The six-row parent                                Correlations between Chemical Traits
Morex was found to carry minor high-protein alleles on
2H, 5H, and 7H.                                                                 Protein content was positively correlated with NDF
                                                                             and ADF content for two-row lines, but not for six-row
                                                                             lines (Table 3). -Glucan content was negatively cor-
    Detection of Additive                 Additive Epistasis                 related with NDF and ADF content for two-row lines,
   The coefficients of variation were low for protein                        but the correlation was inconsistent for six-row lines.
(4.0–4.1%), NDF (3.4–5.6%), and ADF (7.2–8.7%) con-                          NDF content was positively correlated with ADF con-
tent. Significant line    location interactions were de-                     tent for both six-row and two-row barley lines. The posi-
tected for protein, NDF, and ADF content (P 0.01).                           tive correlation between NDF and ADF was expected
Variation among DH lines was significant for all five                        because the former estimates the cell wall components
traits at both locations with the exception of starch con-                   cellulose, hemicellulose, lignin, silica, and heat-damaged
tent at Charlottetown. The mean of the DH lines was                          protein, while the latter estimates lignin and cellulose.
1740                                              CROP SCIENCE, VOL. 41, NOVEMBER–DECEMBER 2001

Protein content was not correlated with -glucan con-                              Cedillo et al., 2000). Three QTL for protein content
tent for either six-row or two-row lines. Therefore, QTL                          and two QTL for -glucan content were reported by
for protein content were not linked with those for                                Hayes et al. (1993) and Han et al. (1995), respectively,
  -glucan content in this cross. This was in contrast with                        on chromosome 2H in the Steptoe/Morex cross. But
what other workers (Hayes et al. 1993; Han et al. 1995)                           none of these QTL are located close to the vrs1 locus.
found in the ‘Steptoe’/‘Morex’ cross. Hayes et al. (1993)                         Therefore, associations between six-row and low protein
identified six QTL for protein content with three on                              and between six-row and low -glucan were most likely
chromosome 2H and one each on chromosome 3H, 4H,                                  due to a pleiotropic effect of the vrs1.a allele. In this
and 5H. Han et al. (1995) found three QTL for barley                              study, six-row lines contained 4% higher starch and 0
  -glucan, one on chromosome 1H and two on chromo-                                to 4% higher NDF than two-row lines. Six-row lines,
some 2H. The ABG703-Chs1B interval on chromosome                                  however, did not differ from two-row lines in ADF
2H was associated with QTL for both protein and -                                 content. Narasimhalu et al. (1995) and Kong et al. (1995)
glucan content. High -glucan content is undesirable                               reported that Canadian two-row cultivars on average
for feed especially for poultry and malt but desirable                            contained less -glucan, more starch, less NDF, and
for human nutrition. Consequently, a lack of correlation                          less ADF than Canadian six-row cultivars. But when
between protein content and -glucan content would                                 comparisons were made with the same average genetic
be a positive attribute to barley breeding programs                               background as in this study, two-row lines contained
which aim at developing barley cultivars with high pro-                           more -glucan, less starch, and similar concentrations
tein and low -glucan content for livestock feeds.                                 of NDF and ADF as six-row lines. The results of this
                                                                                  study showed that the vrs1.t allele had a significant effect
                 Effects of Marker Genes                                          on four of the five chemical traits. Previous studies
                and Chromosome Segments                                           showed that the vrs1.t allele also had an effect on grain
   Two-row lines contained 14 to 20% higher protein                               yield, test weight, seed weight, plant height, lodging
and 6 to 7% higher -glucan than six-row lines (Table 4).                                                                       ´
                                                                                  resistance, seed size, and seed shape (Fregeau-Reid et
The difference between two-row (vrs1.t) and six-row                               al., 1996; Jui et al., 1997). Purple lemma lines contained
(vrs1.a) alleles accounted for 67 to 77% of the total vari-                       10 to 16% higher protein and 5% higher -glucan than
ation for protein content. The results agree with earlier                         yellow lemma lines. In contrast, yellow lemma lines
findings that two-row (vrs1.b) genotypes were associ-                             contained 3% higher starch, 2 to 4% higher NDF, and
ated with higher protein content compared with six-row                            0 to 3% higher ADF than purple lemma lines. Both the
(vrs1.a) genotypes (Day and Dickson, 1957; Pomeranz                               srh and Est1 loci had no effect on the five chemical
et al., 1973; Hockett and Standridge, 1976; Takahashi                             traits. The raw1 allele was associated with high protein
et al., 1976; McGuire and Hockett, 1983; Marquez-                                 content at Ottawa, but it was not associated with starch,

Table 4. Means of 12 classes of the 190 barley doubled-haploid lines grown at Charlottetown (C) and Ottawa (O) in 1993.
                                        Protein                   Starch                     -glycan                 NDF                 ADF
Comparison                        C                O         C              O          C               O        C            O     C           O
                                                                                           g kg
Spike type
  2-row (vrs1.t vrs1.t )        171a†             158a     491a            495a        47a            47a     160a          156a   72a         70a
  6-row (vrs1.a vrs1.a)         150b              132b     510b            514b        44b            43b     161a          162b   72a         70a
  %‡                             67                77       17              18          8              6        0            17     0           0
Lemma color
  Purple (Pre2Pre2 )            168a              155a     494a            496a        46a            46a     159a          156a   72a         69a
  Yellow (pre2pre2 )            153b              134b     507b            513b        44b            44b     162b          163b   72a         71b
  %                              41                59        7              15          6              3        4            24     0           9
Rachilla hair
  Long (SrhSrh )                160a              144a     501a            505a        45a            46a     159a          159a   72a         70a
  Short (srhsrh )               161a              145a     500a            505a        46a            44a     162a          159a   72a         70a
  %                               0                 0        0               0          0              0        0             0     0           0
Awn type
  Rough (Raw1Raw1 )             159a              142a     502a            508a        45a            45a     160a          160a   72a         70a
  Smooth (raw1raw1 )            162a              147b     499a            502a        45a            45a     161a          158a   72a         69a
  %                               0                 4        0               0          0              0        0             0     0           0
Esterase 1
  CaCa                          159a              143a     499a            508a        45a            45a     161a          159a   71a         69a
  PrPr                          161a              143a     503a            504a        43a            45a     162a          159a   73a         70a
  %                               0                 0        0               0          0              0        0             0     0           0
Esterase 5
  MeMe                          161a              144a     496a            504a        46a            45a     163a          158a   73a         69a
  RiRi                          158a              141a     508b            508a        43a            45a     159a          160a   71a         70a
  %                               0                 0        6               0          0              0        0             0     0           0
† Means in each comparison followed by different letters were different at the 0.05 level.
        2   2     2
‡%      M/( M     W)   100%, where 2  M    variance between marker classes and 2     W    variance within marker classes.
                                              FREGEAU-REID ET AL.: CHEMICAL COMPOSITION OF BARLEY                                                                    1741

Table 5. Mean squares for chemical traits of two marker loci on chromosome 2H as identified from 190 barley doubled-haploid lines
  grown at Charlottetown and Ottawa in 1993.
Sources                                           df                   Protein                    Starch                       -glucan              NDF              ADF
  Between classes                                  3                   13 814**                6 044**                     122*                      225             629*
    Spike types                                    1                   40 945**               17 449**                     359**                     628              32
    Lemma colors                                   1                      298                    450                         3                     1 121*            170
    Spike type lemma color                         1                      197                    231                         4                       495             470*
  Lines (classes)                                186†                     257                    887                        38                       275              81
    Lines (vrs1.tVrs1.tPre2Pre2 )                 84‡                     181b§                  586b                       38a                      206b             61b
    Lines (vrs1.tvrs1.tpre2pre2 )                  8                      184ab                  887b                       50a                      165b            106a
    Lines (vrs1.avrs1.aPre2Pre2 )                 11                      119b                   943b                       27a                       54c             43b
    Lines (vrs1.avrs1.apre2pre2 )                 83                      359a                 1 180a                       37a                      385a            105a
  Between classes                                  3                      329                    506                       173*                      277              42
    Spike types                                    1                   63 612**               17 683**                     464**                   3 275**            33
    Lemma color                                    1                      276                    471                         2                     1 343**           646**
    Spike type lemma color                         1                        2                    429                        52                        54             190
  Lines (classes)                                186                      235                    783                        63                       179              68
    Lines (vrs1.tvrs1.tPre2Pre2 )                 84                      249a                   816a                       51a                      178a             66a
    Lines (vrs1.tVrs1.tpre2pre2 )                  8                      244a                   287a                       28a                       65a            123a
    Lines (vrs1.avrs1.aPre2Pre2 )                 11                       92a                   932a                       60a                      184a             82a
    Lines (vrs1.avrs1.apre2pre2 )                 83                      239a                   779a                       78a                      200a             63a
* Indicates significance at P 0.05.
** Indicates significance at P 0.01.
† df 185 for starch and -glucan content at Charlottetown, and for -glucan at Ottawa.
‡ df 83 for starch and -glucan content at Charlottetown, and for -glucan at Ottawa.
§ Variances for the same trait followed by different letters were different at the 0.05 level.

  -glucan, NDF, or ADF content at both locations. Al-                                    either the absence of QTL or the presence of the same
leles at the Est5 locus had a significant effect only on                                 QTL allele in the vrs1-Pre2 segment (Table 5). The four
starch content.                                                                          variances among lines for the four other chemical traits
   The pairwise analysis showed that the vrs1 locus had                                  were homogeneous at Ottawa, but not so at Char-
a significant effect on protein, starch, and -glucan con-                                lottetown (Table 5). These could be due to the presence
tent while the Pre2 locus had a significant effect on NDF                                of line location interactions. Why purple lemma was
and ADF content (Tables 5 and 6). There were no                                          associated with low NDF and low ADF is not known.
interactions between the vrs1 and Pre2 loci for these                                    Purple lemma lines contain anthocyanin pigments (Mul-
chemical traits with the exception of ADF content at                                     lick et al., 1958). The biosynthesis of anthocyanin is
Charlottetown. In single-locus analysis, differences in                                  linked to that of lignin in the shikimic acid pathway
protein, starch, and -glucan content were found to be                                    by that both require p-coumaric acid (Goodwin and
associated with the Pre2 locus (Table 4). These associa-                                 Mercer, 1983). Therefore, competitions for p-coumaric
tions must be caused by linkage between the Pre2 and                                     acid for biosynthesis could lead to reduction of lignin
vrs1 loci, because the pairwise analysis showed little                                   production, which, in turn, could lead to low concentra-
effect of the Pre2 locus on these three chemical traits                                  tions of NDF and ADF in the purple lemma lines. If
(Table 5). Choo (1983) showed that in theory the vari-                                   so, then purple lemma could be used as a selectable
ances of the four marker classes are heterogeneous in                                    marker for low fiber content in barley. Another pairwise
the presence of interactions between QTL and marker                                      analysis showed that the Srh locus, Raw1 locus, and
genes, but become homogeneous when both gene inter-                                      their interactions had no effect on the five chemical
actions and the additive effect of QTL are absent. In                                    traits (Tables 6 and 7). In most cases, the four variances
this study, the four variances among lines for -glucan                                   among lines within the four marker classes were homo-
content were homogeneous at both locations, indicating                                   geneous for these five chemical traits. Therefore, the
Table 6. Means of eight genotypes found in the 190 barley doubled-haploid lines grown at Charlottetown (C) and Ottawa (O) in 1993.
                                                         Protein                     Starch                      -glucan                     NDF                   ADF
Genotype                            # lines        C               O             C            O              C             O             C          O         C          O
                                                                                                      g kg
Spike type and lemma color
  vrs1.tvrs1.tPre2Pre2                85          171a†        158a          491a         494a             47a         46a           160a          156a      72a         69a
  vrs1.tvrs1.tpre2pre2                 9          171a         156a          490a         504ab            47ab        49a           161ab         160ab     71ab        71a
  vrs1avrs1.aPre2Pr2                  12          146b         135b          517b         514b             44ab        44a           154c          156a      67b         65b
  vrs1.avrs1.apre2pre2                84          151b         132b          509b         514b             44b         43a           162b          163b      72a         71a
Rachilla hair and awn type
  SrhSrhRaw1Raw1                      69          159a         143a          503a         506a             45a         45a           159a          160a      71a         70a
  SrhSrhraw1raw1                      30          162a         148b          496a         502a             44a         46a           161a          157a      72a         68a
  srhsrhRaw1Raw1                      27          159a         141a          497a         513a             46a         43a           163a          160a      72a         70a
  srhsrhraw1raw1                      64          162a         147b          501a         501a             45a         45a           161a          159a      72a         70a
† Means in each comparison followed by different letters were different at the 0.05 level.
1742                                            CROP SCIENCE, VOL. 41, NOVEMBER–DECEMBER 2001

Table 7. Mean squares for chemical traits of two marker loci on chromosome 5H as identified from 190 barley doubled-haploid lines
  grown at Charlottetown and Ottawa in 1993.
Sources                                         df                Protein               Starch               -glucan              NDF                ADF
  Between classes                                3                 329                   506                  17                  277                 42
    Rachilla hairs                               1                 197                    77                  39                  460                 82
    Awn types                                    1                 776                   219                  10                    9                 27
    Rachilla hairs awn types                     1                   2                  1223                   0                  367                 22
  Lines (classes)                              186†                474                   976                  17                  281                 84
    Lines (SrhSrhRaw1Raw1 )                     68                 491a§                 861a                 33ab                195a                70a
    Lines (SrhSrhraw1raw1 )                     29                 438a                  998a                 20b                 262a                60a
    Lines (srhsrhRaw1Raw1 )                     26                 384a                  940a                 52a                 439a                98a
    Lines (srhsrhraw1raw1 )                     63‡                511a                 1108a                 50a                 316a               104a
  Between classes                                3                 901                   856                  47                  153                 62
    Rachilla hairs                               1                  76                     1                 105                   67                  1
    Awn types                                    1                2580*                 1971                  36                   14                132
    Rachilla hairs awn types                     1                  46                   597                   0                  379                 52
  Lines (classes)                              186                 564                   869                  65                   65                 71
    Lines (SrhSrhRaw1Raw1 )                     68                 576a                  906a                 56a                 151a                74a
    Lines (SrhSrhraw1raw1 )                     29                 479a                  818a                 53a                 147a                90a
    Lines (srhsrhRaw1Raw1 )                     26                 475a                  781a                108a                 434b                85a
    Lines (srhsrhraw1raw1 )                     63                 627a                  890a                 62a                 185a                54a
* Indicates significance at P 0.05.
** Indicates significance at P 0.01.
† df 185 for starch and -glucan content at Charlottetown, and for -glucan at Ottawa.
‡ df 62 for starch and -glucan content at Charlottetown, and for -glucan at Ottawa.
§ Variances for the same trait followed by different letters were different at the 0.05 level.

results suggested that alleles at both the Srh and Raw1                           ducting the chemical analyses, and L. Langille for conducting
loci had no effect on these traits, and that either no                            statistical analyses.
QTL or the same QTL allele for these chemical traits
were located on the chromosome 5H segment flanked                                                          REFERENCES
by these two marker loci. Hayes et al. (1993) and Han                             AACC. 2000. Approved methods of the American Association of
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  -glucan content, respectively, on this chromosome seg-                             Chemists, Inc., St. Paul, MN.
ment in the Steptoe/Morex cross.                                                  Bouard, J.-P., J. Fekette, and M. Leuillet. 1980. Influence du type
   This study substantiates that DH lines are very useful                            d’orge (2 rangs ou 6 rangs) sur les performances zootechniques du
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                                                                                     porcelet sevre. (In French.) Journees Rech. Porcine en France 12:
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                                                                                  Choo, T.M. 1983. Doubled haploids for locating polygenes. Can.
detected on two chromosome segments.                                                 J. Genet. Cytol. 25:425–429.
                                                                                  Choo, T.M., and E. Reinbergs. 1982. Analyses of skewness and kurto-
                        CONCLUSIONS                                                  sis for detecting gene interaction in a doubled haploid population.
                                                                                     Crop Sci. 22:231–235.
   In conclusion, two-row lines from the Leger/CI9831                             Choo, T.M., K.M. Ho, T. Konishi, and R.A. Martin. 1992. Tests for
cross grown in Eastern Canada contained more protein,                                randomness among doubled-haploid lines derived by the bulbosum
                                                                                     method in barley (Hordeum vulgare L.). SABRAO J. 24:87–92.
less starch, and more -glucan than six-row lines; while                           Choo, T.M., A. Kotecha, E. Reinbergs, L.S.P. Song, and S.O. Fejer.
purple lemma lines contained less NDF and ADF than                                   1986. Diallel analysis of grain yield in barley using doubled-haploid
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                                                                                     Agron. J. 49:244–245.
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content, however, were correlated with NDF and ADF                                   Group Limited, London.
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                                                                                  Fregeau-Reid, J., T.M. Choo, P. Jui, and K.M. Ho. 1996. Inheritance
stasis was detected for starch and NDF content. The                                  of kernel size and shape of barley. SABRAO J. 28:47–55.
results of this study also suggested that selection for                           Goodwin, T.W., and E.I. Mercer. 1983. Introduction to plant biochem-
                                                                                     istry, second edition. Pergamon Press, Oxford, UK.
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                                                                                     and A. Kilian. 1995. Mapping of -glucan content and -glucanase
                     ACKNOWLEDGMENTS                                                 activity loci in barley grain and malt. Theor. Appl. Genet. 91:
   The authors were grateful to Moe Kuc for producing the                         Hayes, P.M., B.H. Liu, S.J. Knapp, F. Chen, B. Jones, T. Blake, J.
DH lines, Sharon ter Beek and Gary Cooper for their technical                        Franckowiak, D. Rasmusson, M. Sorrells, S. E. Ullrich, D. Wesen-
assistance in conducting the field trials, E. Imbeault for con-                      berg, and A. Kleinhofs. 1993. Quantitative trait locus effects and
                            LUKASZEWSKI ET AL.: TRANSFER OF RWA RESISTANCE FROM A RYE CHROMOSOME                                              1743

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     Attempts to Transfer Russian Wheat Aphid Resistance from a Rye Chromosome
                             in Russian Triticales to Wheat
         Adam J. Lukaszewski,* David R. Porter, Cheryl A. Baker, Krystyna Rybka, and Boguslaw Lapinski

    The Russian wheat aphid (Diuraphis noxia Mordvilko) is a serious
pest of wheat (Triticum aestivum L.). To extend the range of genetic
                                                                             E     xtensive crop damage caused by the Russian wheat
                                                                                   aphid (RWA) (Diuraphis noxia Mordvilko), first
                                                                             in South Africa and later in the United States, spurred
variation of resistance, attempts were undertaken to transfer near-          energetic efforts to identify sources of resistance and to
immunity to RWA into wheat from two Russian triticales (X. Tritico-
                                                                             incorporate the appropriate loci into wheat (Triticum
secale Wittmack) PI 386146 and PI 386156 by irradiation and by
induced homologous recombination. The rye genome in the triticale
                                                                             aestivum L.) cultivars. Screening of gene bank acces-
lines was derived from Secale montanum Guss. Tests of resistance in          sions identified numerous exotic wheat lines as well as
early backcrosses to wheat indicated that the near-immunity of the           some related species with various levels of resistance
triticale lines was controlled by at least two loci, one of which was        (du Toit, 1985, 1987; Nkongolo et al., 1989; Porter et al.,
located on rye chromosome arm 4RLmon. Centric wheat-rye transloca-           1993). Among wheat relatives, high levels of resistance
tion 7DS.4RLmon that appeared to be compensating, was produced.              were observed in rye (Secale cereale L.) with near-immu-
To further reduce the amount of rye chromatin present, its long arm          nity present in a group of four triticales (X. Triticosecale
was induced to recombine with wheat chromosomes by the removal               Wittmack), PI 386148, PI 386149, PI 386150, and PI
of the Ph1 locus. Among 3563 progeny screened, only two wheat-rye            386156, from the N.I. Vavilov All-Russian Scientific Re-
recombinant chromosomes were recovered. Both appeared to be non-
                                                                             search (VIR) Institute of Plant Genetic Resources in
compensating and were involved in multivalents in meiosis. Irradiation
of PI 386156 followed by crosses and backcrosses to wheat with several
                                                                             the Russian Federation (Nkongolo et al., 1989; Webster,
generations of selection for resistance resulted in a wheat line that        1990). Results from a genetic study of resistance in three
was found to be a disomic addition of chromosome 4Rmon–centric               of these triticales (PI 386148, PI 386149, and PI 386156)
translocation homozygote of rye chromosome tentatively identified            were interpreted as indicative of a single gene mode of
as 5Rmon. With only one locus for resistance from the original triticale     inheritance (Nkongolo et al., 1992). However, a detailed
parents, the addition line of 4Rmon, centric translocation line 7DS.4RLmon   study of PI386156 by Fritz et al. (1999) disputed the
and recombinant lines of 4RLmon had only moderate level of resistance        single-gene control of resistance. Also, PI 386148 and
to RWA. The study demonstrates that transfers of alien variation             PI 386156 showed different reactions when challenged
into wheat may be severely complicated by unclear genetics of the
                                                                             by a virulent French RWA isolate, indicating these two
target traits, low levels of homology, and structural differences be-
tween the donor and recipient chromosomes.
                                                                             triticales have different RWA resistance mechanisms
                                                                             (Puterka et al., 1992) while PI 386148 and PI 386150
A.J. Lukaszewski, Dep. of Botany and Plant Sciences, University of
                                                                             showed different levels of resistance in the study of
California, Riverside, CA 92521; D.R. Porter and C.A. Baker, USDA-           Nkongolo et al. (1996).
ARS, 1301 N. Western Rd., Stillwater, OK 74075-2714; K. Rybka and               The levels of resistance of the four Russian triticales
B. Lapinski, Institute of Plant Breeding and Acclimatization, 05-870         to the RWA population in the USA are much higher
Blonie, Poland. Received 16 Jan. 2001. *Corresponding author (ajoel@         than those found in hexaploid wheats (Webster, 1990)
                                                                             and would be beneficial in developing new RWA-resis-
Published in Crop Sci. 41:1743–1749 (2001).                                  tant wheat cultivars. This article describes attempts to

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