**9. Genomics-assisted breeding (GAB)**

*Legume Crops - Prospects, Production and Uses*

**8. Breeding for amino acid composition**

acids was not changed compared to parental lines.

were identified using induced mutation.

primer pairs specific to *Gy1*, *Gy2*, and *Gy5* were designed [93–95]. PCR primers were designed for identification of the *Gy4* null allele and demonstrated selection of soybean without the A4 peptide can be done by null allele specific primers which overcome the drawbacks of SDS-PAGE gel-based selection for A4 peptide [95].

Besides breeding for increased protein content, protein composition is important for its nutritional value. Based on solubility properties, globulins and albumins are two major components of dicot seed storage protein, and soybean primarily belongs to the globulin (~70%) family [96]. The soybean globulins (glycinin and β-conglycinin) are relatively low in sulfur-containing amino acids methionine (Met) and cysteine (Cys) as well as threonine (Thr) and lysine (Lys) [97]. Increasing the soybean storage protein content of seed along with improving the ratio of glycinin to β-conglycinin is of great potential for food grade soybean improvement [98, 99]. Therefore, besides increased protein content, enhancing sulfur containing amino acids (Met, Thr, Cys, and Lys) would improve the nutritional value. More than 70% of the essential amino acid enriched meal is used in the feed industry [97, 100]. Although soybean cultivars with improved protein content have been successfully developed, only a few studies have been conducted to identify genomic regions controlling amino acid composition. The difficulty in breeding for improved amino acids could be due to lack of genetic variability, lack of high throughput, and cost-effective phenotyping platform to screen a large number of samples for amino acids. Panthee et al. [99] identified QTL for essential amino acids in a F6-derived recombinant inbred population. In another study, a major QTL for essential amino acids and crude protein was identified on Chr20 [97]. Moreover, negative correlations of crude protein with Lys and Thr and a positive correlation between Thr with Lys were also observed [97]. Among the essential amino acids, Met, Lys, and Thr are synthesized from a common precursor aspartate; thus, they are strongly correlated. Krishnan et al. [101] introgressed leginsulin (Cys-rich protein) and a high protein trait from an Asian soybean germplasm, PI 427138, into North American experimental line (LD00–3309). While they were successful in introgressing leginsulin and improving protein content, the overall concentration of sulfur-containing amino

Seed protein content and composition are dependent on the genetic background of an elite parent that plays an important role in the expression of a newly introgressed allele because of complex epistatic interactions [102]. It has been found that most of the QTLs affecting seed protein and yield and yield-related components were detectable only in one of the parental genetic backgrounds (GBs) in introgression lines of reciprocal crosses [103]. The high protein allele within a different genetic background resulted into reduced Thr and Lys content [103]. The high protein allele from Danbaekkong on Chr20 has been demonstrated to increase seed protein content in several maturity groups (III–VIII) in various genetic backgrounds with little drag on seed yield [104]. On the other hand, yield drag was observed for the protein QTL alleles on Chr20 from other sources, including wild *G. soja* [56, 60]. Hence, it is not feasible to select only the major crude protein QTL on Chr20 to improve protein quality. Improvement of protein and amino acid profiles has been limited by the narrow genetic base and genome complexity of soybean. Mutation breeding can be used to enhance the genetic variability. Mutagenized populations (physical, chemical, transposon tagging or transformation-induced mutagens) have been useful in crop improvement [105]. In soybean, mutations for seed traits, including oleic acid [106], oil [105, 107], stearic acid [108], and lipoxygenase [109]

**62**

The integration of genomic tools and breeding practices are the core components of genomics-assisted breeding (GAB) for developing improved cultivars for any given trait. Near-isogenic lines (NILs) can be developed for major QTL (e.g., protein QTL on Chr20) by backcross breeding. Using NILs, the effect of a QTL and the phenotype it produces (i.e., protein or amino acid content) can be estimated precisely without the confounding effects of differences in genetic backgrounds. Additionally, developing NILs in a range of maturity groups is desirable to study the effect of environment and maturity on seed protein content. Marker-assisted backcrossing selection approach was utilized to produce a NIL-(cgy-2–NIL)-containing mutant cgy-2 allele, responsible for the absence of allergenic α-subunit of β-conglycinin [110]. It is also possible to incorporate multiple genes/QTL into elite lines in a cyclic forward crossing scheme and employing marker-assisted recurrent selection (MARS) as an effective approach [111, 112]. Recurrent selection was effectively utilized for increased gain yield, protein, oil, and oleic acid content [111, 113, 114]. Furthermore, the next-generation sequencing (NGS) data can be used effectively for genomic selection (GS) to identify desirable parents and progenies. Jarquin et al. [115] assessed the genomic and phenotypic data of over 9000 accessions and developed genomic predication models to evaluate the genetic value for protein, oil, and yield traits. Similarly, genomics-assisted haplotype analysis is a promising approach if the information of a major QTL is available and that can be applied to select desirable haplotype blocks for parental selection and crossing by design [116].

In order to widen the genetic base, it may be necessary to utilize wild species accessions as introgression libraries as well as developing interspecific populations. On the other hand, elite cultivars and landraces can be used to develop mapping populations, and training populations [114]. Wild soybean (*G. soja*) serves as a unique resource to study regulation of protein and amino acid biosynthesis, because the seed concentration of these components is higher in *G. soja* compared with *G. max.* Utilization of *G. soja* in breeding program is hampered due to linkage drag on favorable agronomic characteristics [113]. However, this issue could be resolved by advanced backcross QTL-based breeding, which was utilized for introgressing alleles from wild tomato to cultivated type for yield improvement [117], or through mutation breeding approaches.
