BGN 19: A Proposal on identification of major genes in spring barley using morphological markers

A Proposal on identification of major genes in spring barley using morphological markers

R.D. Horsley and J.D. Franckowiak
Crop and Weed Sciences Department
North Dakota State University
Fargo, North Dakota 58105, U.S.A.


The goal of the barley breeder is to develop new cultivars that are superior to the "best" existing cultivars. This is accomplished usually by combining large numbers of favorable genes for agronomic and malt quality. Many of the gains made in plant breeding are relatively small, but they accumulate over time and result in the frequent release of improved cultivars. Thus, much money and time is invested to identify new cultivars.

To increase the probability and efficiency of obtaining superior new cultivars, it would be advantageous to identify genes that have a large effect on agronomic traits and malt quality. Most of the economically important traits of barley (e.g. yield, test weight, and many malt quality parameters) are known as quantitative or polygenic traits. However, not all traits of interest are controlled by this type of gene action. In animals, many traits that are generally regarded as polygenic have been associated with a single gene (Crawford and Smith, 1964; Hartman, 1972; Merat, 1986; Pirchner, 1988). A major gene affecting a quantitative trait that has been localized to a chromosome is called a quantitative trait locus (QTL) by Gelderman (1975). If QTLs can be identified in barley, they can be evaluated and accumulated in improved lines.

Two methods of identifying QTLs are available currently. The first method involves utilization of restriction fragment length polymorphisms (RFLPs), isozymes, and other molecular markers. Utilization of molecular marker techniques requires special facilities, equipment, and personnel; thus, these methods are expensive and unavailable to many barley breeders. The second method is to identify QTL linked to a morphological marker locus. Morphological markers are simply inherited genes that cause a visual change in a morphological trait such as kernel color, surface waxes, plant height, etc. Because a large number of morphological markers have been mapped, this method is available to barley breeders and does not require special equipment, facilities, or personnel. A proposal utilizing morphological marker loci to identify QTLs is outlined and discussed below.

Procedure :

After suitable morphological markers are selected and placed in a common genetic background, identification of a QTL is a four step process. First, a possible association between a QTL and morphological marker locus must be identified. Secondly, it must be determined if the association between the morphological marker locus and the proposed QTL is due to linkage or pleiotropy. Next, if linkage is observed, the QTL needs to be localized in relation to other morphological markers on that chromosome. Finally, it must be determined if the QTL is omnipresent in elite breeding material.

Identification of Possible QTL Associations

BC1 lines can be used to identify possible associations between a morphological marker locus and QTL. The recurrent parent should have excellent agronomic and malt quality and the donor parent with the morphological marker should have relatively poor agronomic and malt quality as compared to the recurrent parent.

Agronomic quality of BC1F3 and BC1F lines can be evaluated in yield trial experiments conducted over two summers at two locations. Growing the experiments over two years will allow an evaluation of the genotype X environment effects. An augmented block design (Federer, 1961) should be the best experimental design to detect significant differences between BC1 lines and other entries in the experiment. Use of an augmented block design would be advantageous over replicated designs for several reasons. Replicated yield trials would require an extra generation to increase seed for yield trials and would occupy more area in the field. Also, because of the greater size of replicated yield trials, only five to ten morphological marker loci could be evaluated each year for possible associations to QTLs. With an augmented block design, up to 50 morphological marker loci could be screened each year. For midwestern six-rowed breeding material, entries that could be included in each block are the recurrent parent, Morex as the quality check, the donor parent, and seven to ten BC1F3 lines having the same morphological marker.

Field notes should be taken on all desired agronomic traits (e.g. heading date, plant height, foliar diseases, lodging, etc.) At maturity, each plot would be harvested and individually threshed. Data to be collected after harvest could include grain yield, test weight, 1000- kernel weight, percent barley protein, kernel assortment and other characters of interest. Malt quality can be evaluated on samples of grain obtained from the yield trial experiments. Malt parameters to be studied could include percent soluble wort protein, percent fine-grind extract, percent coarse-grind extract, fine-coarse grind differences, diastatic power, alpha-amylase activity and other parameters of interest.

Possible associations between a morphological marker locus and a QTL can be determined statistically. If BC1 progeny are distributed normally between the donor line and the recurrent parent, no association between the marker locus and a QTL for a specific trait should be assumed. An association between a marker locus and a QTL is possible if the values for BC1 lines are skewed towards the donor line.

Determination of Linkage vs. Pleiotropy

For those traits showing an association with a genetic marker, it will be necessary to determine if the association is due to pleiotropy or due to tight linkage with a QTL. To screen for linkage, a BC1F4 line that has a low value for the trait in question and expresses the morphological marker should be backcrossed to the recurrent parent. Two spikes from 1,000 randomly selected plants should be harvested from the BC2F2 population. The progeny should not be selected for expression or non- expression of the morphological marker. BC2F2 derived BC2F3 families should be grown in head rows and notes should be taken on the morphological marker and the trait in question for each family. Presence of linkage can be determined using chi-square analysis. Fisher's Maximum Likelihood Method can be used to estimate linkage intensity between the marker locus and the QTL.

Evaluating if a QTL is Omnipresent

Other elite breeding material should be evaluated for the presence of the QTL. To screen the germplasm, a BC1F4 line that is a recombinant (i.e. a BC1F4 line with the marker locus and quality resembling the recurrent parent for the trait in question) can be crossed to several elite lines. In the F2 generation, two spikes from at least 50 plants expressing the marker phenotype should be selected. F2 derived F3 families should be grown in head rows and notes should be taken on the trait(s) in question for each family row. The QTL will be deemed absent in the elite line if transgressive segregates are found in the F2 derived F3 families. If all families appear similar, then the same QTL will be assumed to be present in both populations.


Ideally, morphological markers and molecular biology techniques would be combined to develop a map of the barley genome which includes morphological marker loci, enzymatic loci, RFLPs, and QTLs. With such a map, even barley breeders having limited resources could manipulate important QTLs. Several researchers (Tanksley et al., 1981; and Tanksley et al., 1982) have shown that introgression of a favorable QTL from unadapted material is possible. Selection for the marker along with the linked QTL will reduce the generations required to complete backcrossing and space required for evaluation of segregating progeny.

Possible associations between morphological marker loci and QTLs have been studied at North Dakota State University (Horsley, 1988; Gonzalez, 1990). Horsley identified a possible association between the ant13 gene of DM582 on chromosome 6 conditioning for low polyphenol content (Falk, 1985; Hormis, 1988) and a major gene for diastatic power (DP). Gonzalez (1990) identified a possible association between the v3 gene on chromosome 5 for intermediate six-row type (Fukuyama, 1983) and a major gene conditioning resistance to spot blotch, incited by Cochliobulus sativus (Ito and Kurib.) Dreschs. ex Dast.

Continued research on the ant13 gene will include determining if the association is due to linkage or pleiotropy. If the association is due to linkage, linkage intensity between the QTL affecting DP and the ant13 gene will be studied. Since the ant13 has been located on chromosome 6, linkage intensity will be determined between the QTL for DP and other morphological marker genes located on chromosome 6. If the association is due to linkage, it will be determined if the QTL affecting DP is present in barley lines having inherently low DP. If it is not, this QTL could be introgressed into such lines to further increase DP.


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