Soybean - Bioenergy Crop Plants - page 514

20.8 lInolenIc acId
Generally soybean oil has 8-10% linolenic acid (Wilson 2004). Linolenic acid (18:3) has been iden-
tified as an unstable component of soybean oil (Liu and White 1992). One of the most important
goals of oil quality breeding in soybean has been to reduce its linolenic acid content to improve
oxidative stability and flavor and to reduce the need for hydrogenation. Several linolenic acid altered
genotypes were developed by recurrent selection, mutagenesis, and germplasm screening.
Through several cycles of recurrent selection, the N79-2245 soybean with 51% oleic acid and
4.2% linolenic acid was developed (Wilson et al. 1981). At least three independent genetic loci
are associated with seed linolenic acid levels, with mutant alleles identified at fan , fan2 , fan3 , and
fanx . Also, multiple alleles at the fan locus have been reported. Mutant C1640 ( fan , 3.4% 18:3)
(Wilcox et al. 1984; Wilcox and Cavins 1985), A5 ( fan , 4% 18:3) (Hammond and Fehr 1983a;
Rennie and Tanner 1991), A23 ( fan2 , 5.6% 18:3) (Fehr et al. 1992), KL-8 ( fanx ) and M-5 ( fan )
(Rahman et al. 1996b), M-24 ( fanx a ) (Rahman et al. 1998), and RG10 ( fan-b , < 2.5%) (Stojsin et
al. 1998) were developed by mutagenesis. The fan alleles were also found in the USDA Soybean
Germplasm Collection. PI123440 and PI361088B contained natural mutations at the Fan locus that
have been shown to be either allelic or identical to the original fan allele in C1640 (Howell et al.
1972; Rennie et al. 1988; Rennie and Tanner 1989a). Evidence of two loci ( fanfan2 ) were found in
A16 and A17 (Fehr et al. 1992), MOLL ( fanfanx ) (Rahman and Takagi 1997), and LOLL ( fanfanx a )
(Rahman et al. 2001). A29, a very low (1%) linolenic acid line, was reported (Ross et al. 2000). A29
was developed by combining three independent mutations: fan from line A5, fan2 from A23, and
fan3 from a mutagenized derivative of line A89-144003 (Ross et al. 2000).
The discovery of the molecular genetic basis for the low linolenic acid trait in soybeans was based
on using gene information from experiments done in Arabidopsis spp. and mutagenized soybean
lines identified with the low linolenic acid trait. In Arabidopsis spp., it was discovered that a single
gene, FAD3 , controlled the conversion of linoleic acid precursors into linolenic acid precursors in
the seed oil (Yadav et al. 1993). The soybean genome was shown to have at least three versions of
the FAD3 gene (Bilyeu et al. 2003; Anai et al. 2005). By sequencing and characterizing the FAD3
alleles in soybean lines with low linolenic acid, corresponding mutations could be found in three
of the genes that affected linolenic acid content (Bilyeu et al. 2003, 2005, 2006; Anai et al. 2005;
Chappell and Bilyeu 2006, 2007). Because the sequence changes identified in the alleles defined the
mutations, perfect molecular markers were designed to specifically select the desired alleles (Bilyeu
et al. 2005, 2006; Chappell and Bilyeu 2006, 2007). Selection by breeder-friendly perfect molecular
markers can be an efficient system for soybean improvement, particularly if a backcross strategy is
used (Beuselinck et al. 2006; Bilyeu et al. 2006).
Molecular markers closely associated with linolenic acid alleles have been found (Table 20.2).
The Fan locus has been mapped to Chro. 14 (LG B2) of soybean (Brummer et al. 1995). Four RFLP
markers were found using a population from the cross C1640 × PI479750 and were successfully
anchored to markers mapping the Linolen1-2 QTL on Chro. 14 (LG B2) (Diers and Shoemaker
1992; Brummer et al. 1995). In another study, a DNA fragment missing from the ω3 desaturase gene
in soybean line A5 was mapped to the same region of Chro. 14 (LG B2) as the Fan locus, suggest-
ing that a deletion in this gene was responsible for the reduction in 18:3 in A5 (Byrum et al. 1997;
Pantalone et al. 2004).
A study identified and characterized three soybean microsomal ω3 fatty acid desaturase genes—
GmFAD3A , GmFAD3B , and GmFAD3C —and determined that the deletion of the GmFAD3A gene
is responsible for the reduced 18:3 in line A5 (Bilyeu et al. 2003). A point mutation was detected in
the GmFAD3A gene in mutant C1640 at nucleotide 798 of the coding sequence. A guanine residue
was changed to an adenine residue in the C1640 allele, resulting in a change from a tryptophan
codon to a premature stop codon. A molecular marker assay based on a PCR reaction was devel-
oped to distinguish between wild-type Williams 82 and low 18:3 line C1640 alleles (Chappell and
Bilyeu 2006).