Inheritance of Seed Protein Subunits of Common Buckwheat (Fagopyrum esculentum Moench) Cultivar Sobano and Its Homostylous Wild Type

The seed protein subunits of common buckwheat Sobano (Fagopyrum esculentum) and its homostylous wild type “HOMO” (F.esculentum var. homotropicum) were studied by means of SDS-PAGE analysis. The results showed that there are 33 bands of seed protein subunits in Sobano and 31 bands in HOMO. Most of them were found in the cotyledon and a few in the endosperm. There are nine subunits in Sobano seeds and ten subunits in HOMO. The inheritance analysis of the seed protein subunits in progenies of hybrids between Sobano and HOMO showed that there are four pairs of protein subunits controlled by a single co-dominant allele. The genetic analysis of morphological characters indicated that homostyly, shattering and acute achenes are controlled by different dominant single genes. The shattering gene has a linkage to the homostyly locus in a rate of 7.81% and to the allele PS44/PS42.9 in a rate of 22.54%.


Introduction
Common buckwheat (Fagopyrum esculentum) is the key domesticated species mainly grown in the temperate zones of the northern hemisphere: Europe, China, North America, Korea and Japan (Campbell, 1997).The homostylous wild type is close relative to common buckwheat distributed in the Yunnan and Sichuan provinces of China (Chen, 2012).The buckwheat fruit is an achene consisting of an endosperm and an embryo being covered by a testa and a pericarp.The protein content ranges from 8.51 to 18.87 % depending on the cultivar (Krkoskova & Mrazova, 2005).The storage protein prolamin and glutelin contents of buckwheat, however, are very low.Albumin and globulin are the major storage proteins in buckwheat seeds (Guo & Yao, 2006;Tang & Wang, 2010).The 13S salt soluble globulin is the most important one (Radovic et al., 1996;Khan et al., 2012).One of the 13S globulin fractions is expressed within the range of 32-43 kDa, the other between 23-25 kDa (Maksimovic et al. 1996).Buckwheat proteins are characterized by their high nutritional and biological values (Bonafaccia et al., 2003).
The aim of the present study is the identification and characterization of the seed protein subunits of the common buckwheat cultivar Sobano, its wild type and the inheritance of several genes in the progenies of hybrids between them.A further objective is the inheritance of the morphological traits homostyly, shattering, and acute achene.

Plant Material and Crossing Plan
Seeds of the common buckwheat cultivar Sobano possessing dimorphic flowers (heterostyle) and the wild type HOMO with homomorphic flowers (homostyle) were kindly provided by South-West Seed Company (SWS)

Protein
The spectr illustrated of the com respectivel protein sub

Allel
The

The Inheritance of Morphological Characters
All five plants of the F 1 hybrids between Sobano and HOMO showed homostyly, acute achenes, and the shattering habit, indicating a dominant mode of inheritance.Segregations of the three morphological characters in F 2 progenies are listed in Table 3.It is clear that all three pairs of alleles (homostyle/heterostyle, shattering /non-shattering and acute/blunt achene fit a ratio of 3:1, indicating the inheritance model of one pair of alleles.The linkage analyses of the morphological characters are listed in Table 4.The segregation of homostyly and shattering habit do not fit the ratio of 9:3:3:1, indicating a linkage relationship of 7.81% (=1-2× 17/80 ).Since the segregation of the two pairs of relative characters and the achene (acute / blunt) fit the ratio of 9:3:3:1, they are all independent of the achene character (acute / blunt).

The Linkage Analysis of Morphological Characters and Seed Protein Subunits
The segregation of morphological characters and seed protein subunits in 80 F 2 progenies inferred from F 3 lines are listed in Table 5.Only the segregation of PS 44 /PS 42.9 and shattering habit does not fit the ratio of 3:6:3:1:2:1, indicating a linkage relationship of 22.54% (=1-2 12/80 ).Krkoskova and Mrazova (2005) reported that the protein content in buckwheat seeds ranges from 8 19 %, mainly distributed in the aleurone layer and the cotyledon.This study showed that most of seed protein subunits in Sobano and HOMO are distributed in the cotyledon and in endosperm, which is consistent with Maksimovic et al. (1996).Zeller et al. (2004) revealed that two pairs of seed protein subunits (20-25 kD) in the globulin fraction of the seeds fit a co-dominant monogenic mode of inheritance.The present study showed the inheritance of the protein subunits 37-62 kD in the cotyledon of the seeds and found four alleles controlling four pairs of protein subunits controlled by a single co-dominant allele.It further found the inheritance of the protein subunits (37-62 kD) in the cotyledons of the seeds and detected four alleles controlling four pairs of seed protein subunits fitting a co-dominant and single gene pattern in different linkage groups.
In the present study, a one-gene model is postulated for the inheritance of self-compatibility and flower morphology when common buckwheat is crossed with the wild HOMO.Woo et al. (1997), Campbell (1998), Wang and Campbell (1998) and Zeller and Hsam (2001) also proposed the same model for the inheritance of flower morphology.The homomorphic flower type of HOMO is expressed by the allele S h .The complex in cultivated F. esculentum is governed by a single locus S with two alleles S and s that control the reaction in the two types of heteromorphic plants.The relationships between the alleles which produce the three types of flours were described as S > S h >s.Aii et al. (1998) have identified a RAPD marker tightly linked to the S allele at a distance of 0.6 cM.Wang et al. (2005b), however, suggest a two complementary dominant gene loci model in which the loci S h and S c control self-compatibility in HOMO with three alleles at the first locus S h and two alleles at the second locus S c .
In the present study a single dominant gene was found being responsible for seed shattering confirming Ohnishi (1999) who detected in F.esculentum ssp.ancestrale, the ancestor of common buckwheat, also governing by single locus .Matsui et al. (2003), however, using self-compatible lines derived from an interspecific cross between cultivated F. esculentum and the wild type HOMO found that the shattering habit is controlled by two complementary dominant genes.Five AFLP markers were linked to one locus (sht1) and two of these markers co-segregated with the shattering locus without recombination (Matsui et al., 2004).Wang et al. (2005a) postulated for the inheritance of the shattering character the presence of three complementary dominant genes.In the present study a linkage of 7.81% between the genes for shattering and homostyly was observed confirming a previous report by Fesenko et al. (1998).Furthermore this study showed a linkage between the allele pair of shattering and the locus for the seed protein subunits PS 44 /PS 42.9 of 22.54%.
The trait acute/blunt achenes, to our knowledge for the first time described, follows a dominant mode of inheritance being controlled by a dominant gene.
Since all of F 2 progenies inferred from F 3 lines produced by selfing of F 2 plants being homostyly were homostyly, we could not obtain the exchange value of homostyly with PS39.9/PS37.8.It is clear, however, that there is a linkage group of the three loci for shattering habit, homostyly and the protein subunits PS44/ PS 42.9 and the exchange rate of PS 42 /PS 42.9 and homostyly (H/s) (%) is 22.54% + / -7.81% = 30.35%/ 14.73%.Chen et al. (2007) reported that the s gene for long style is located on buckwheat chromosome 4E by means of trisomic lines.
According to the above analysis, there may be six linkage groups discovered in this study, that is, a linkage group of three alleles: shattering, homostyly, and protein subunits PS44 / 42.9 and four linkage groups covering three alleles of seed protein subunits and one locus for the acute achene character, respectively.

Table 1 .
Segregation of four pairs of protein subunits in 63 F 2 progenies (F 2 ) and 80 F 2 progenies inferred from F 3 lines (F 2 *)

Table 2 .
The segregation of two protein subunit alleles in 63 F 2 progenies (F 2 ) and 80 F 2 progenies inferred from F 3 lines (F 2 *)

Table 3 .
Segregation of three morphological characters in 80 F 2 progenies inferred from F 3 lines (F 2 *)

Table 4 .
Segregation of three pairs of relative characters in F 2 progenies

Table 5 .
The segregation of morphological characters and seed protein subunits in 80 F 2 progenies inferred from F 3 lines (F 2 *)