Carrots are grown and consumed by people worldwide and are one of the dietary staples in the United States, China and Indonesia. Carrots are valuable for their taste, good digestibility, and high contents of provitamin A, other carotenoids, and fibers. The popularity of carrots has also been influenced by the introduction of the convenient prepackaged "cut & peel" or "baby carrots," making carrots a leading vegetable snack item.
Biotechnological applications in carrot improvement
The genetics of two multigenic traits of carrot, carotenoids and restoration of CMS were evaluated in the 1960s-1980s. Several individual genes were and the heritabilities of several carrot traits have been measured. Multigene trait mapping only began with the development of molecular markers and linkage maps. To date, seven monogenic traits have been mapped for carrot: yel, cola, Rs, Mj-1, Y, Y2 and P1. Work is underway to localize genes for flavour compounds and pigments, and to localize genes to specific chromosomes using fluorescent in situ methods. QTL have been mapped for carrot total carotenoids and five component carotenoids; phytoene, a-carotene, Beta-carotene, zeta-carotene, and lycopene (Santos and Simon, 2002) and the majority of the structural genes of the carotenoid pathway is now placed into this map (Just et al., 2007). Marker-assisted selection has been reported for Rs gene (Yau et al., 2005).
Bradeen and Simon (1998) studied 103 F2 individuals of the cross B9304 X YC7262, which segregated for core color. Using bulked segregant analysis combined with F2 mapping, they identified six AFLP markers linked to and flanking the Y2 locus. Markers were located between 3.8 and 15.8 cM from the gene. Using the same F2 mapping population, Vivek and Simon (1999) subsequently identified a single AFLP marker located 2.2 cM from the Y2 locus, assigning the locus to one end of linkage group B. Anthocyanin accumulation in the carrot phloem is conditioned by the P1 locus, with purple (P1) dominant to nonpurple (p1) (Simon, 1996). Simon (1996) studied the inheritance of P1 and Y2 in F2 and BC populations originating from Eastern carrot germplasm and concluded that the two loci are unlinked. Consistent with this, Vivek and Simon (1999) mapped P1 to linkage group A, independent of Y2. P1 is flanked by AFLP markers mapping 1.7 and 8.1 cM away from the gene.
The rs allele has recently been found to be a naturally occurring knockout mutant of a carrot invertase isozyme which produces no functional enzyme (Yau and Simon 2003). Vivek and Simon (1999) mapped Rs to one end of linkage group C, 8.1 cM away from an AFLP marker. Mapping results are consistent with inheritance data indicating that Rs is genetically unlinked to Y2 and P1 (Simon, 1996).
Quantitative Trait Loci (QTL) Detected
In addition to single genes conditioning important traits, several QTL have been identified in carrot through segregation analysis. To date, QTL conditioning synthesis in carrot roots of provitamin A a-and Beta-carotenes, the carotene lycopene, and precursors in the carotene pathway have been mapped. Among orange carrots, heritability of 0.40 and approximately 20 major QTL have been reported to control carotenoid content (Santos and Simon, 2002). Most modern carrot breeding effort has exclusively involved intercrosses among orange carrots and the numerous QTL involved in that color class. A major exception to this generalization has been the use of white wild carrot as a source of CMS. As yellow, red, and even white cultivated carrots become more popular, the major genes and eventually QTL conditioning these colors will be better described.
Just et al. (2009) reported that two major interacting loci, Y and Y2 on linkage groups 2 and 5, respectively, control much variation for carotenoid accumulation in carrot roots. These two QTLs are associated with carotenoid biosynthetic genes zeaxanthin epoxidase and carotene hydroxylase and carotenoid dioxygenase gene family members as positional candidate genes. Dominant Y allele inhibits carotenoid accumulation. When Y is homozygous recessive, carotenoids that accumulate are either only xanthophylls in Y2__ plants, or both carotenes and xanthophylls, in y2y2 plants. These two genes played a major role in carrot domestication and account for the significant role that modern carrot plays in vitamin A nutrition.
Few molecular markers in or linked to carrot major genes or QTL have been developed. Examples have been reported for carotene QTL (Santos and Simon, 2002) and the Y2 gene (Bradeen and Simon, 1998) and the Rs sugar type gene (Yau and Simon, 2003; Yau et al., 2005), with marker-assisted selection exercised successfully in the latter case. Yau et al. (2005) suggest that these identifications can be done on leaf tissue from early growth, thereby removing the need for growing mature roots and analyzing sugar content. The identification of markers for soluble solids, carotenoids, and tocopherol, should be feasible and useful as the genetics of these important traits are studied. As codominant markers are more widely developed and maps are joined, the application of these genomic tools can have immediate application in marker-based breeding.
Marker-assisted selection has only recently been initiated in carrot. As the genetics of important complex traits become described, markers to more efficiently select them will warrant development. An international collaborative effort to establish robust genomic platforms (e.g. mapping populations, expression sequence tags, subtractive and large-insert libraries, allele mining, large-scale sequencing banks and expression arrays) for carrot would greatly facilitate the identification of molecular markers and discovery of genes associated with traits of breeding interest.
Among cloned genes with obvious agricultural significance are genes conditioning carbohydrate accumulation in the root. Two enzymes are responsible for the conversion of sucrose to simple sugars. Sucrose synthase catalyzes the reversible conversion of sucrose to UDP-glucose and fructose. The enzyme also plays a role in starch and cellulose biosynthesis and other cellular activities. The gene appears to play an important role in plant growth and is potentially a major determinant of carrot yield. Invertase catalyzes the irreversible cleavage of sucrose to glucose and fructose. Thus, invertase is a major determinant of carrot flavor. Genes encoding both enzymes have been isolated from carrot and subsequently characterized in planta.
Two sucrose synthase genes, Susy1*Dc1 and Susy1*Dc2 have been isolated from carrot. These two sucrose synthase genes differs markedly in their expression patterns. Northern analyses revealed that Susy1*Dc1 is expressed in leaves, roots, flowers, and developing seeds, but Susy1*Dc2 is expressed exclusively in carrot flowers. Several experiments have concluded that sucrose synthase appears to play an important role in metabolic activities associated with plant growth, however it is not significantly involved in sucrose partitioning in carrot.
A direct role of soluble acid invertase in sucrose partitioning and flavor development in the carrot root was established by Yau and Simon (2003) using a candidate gene approach. The dominant Rs allele conditions accumulation in carrot roots of the simple hexose sugars, fructose and glucose. Homozygous rs/rs individuals accumulate sucrose in the roots. The Rs locus is a key determinant of carrot root sugars and flavor. Towards characterization of the molecular basis of the Rs locus, Yau and Simon (2003) generated near isogenic Rs/Rs and rs/rs lines from the carrot inbred B4367. They used RT-PCR to detect transcripts of key enzymes in the sucrose/simple sugar pathway including sucrose synthase, extracellular acid invertase, and two isoforms (designated isoform I and II) of soluble acid invertase. They observed a perfect linkage between the genetically defined Rs locus and the observed mutant soluble acid invertase isoform II allele, providing strong evidence that the Rs locus encodes for soluble acid invertase isoform II.
Genes conditioning root pigmentation and sugar and terpenoid content are candidates for gene mapping in the near future. MAS may be more widely integrated into carrot breeding programs as converted markers linked to quality traits are generated. Genomics resources such as expressed sequence tag libraries, microarrays, in situ hybridization methodologies, and transposon-tagging systems are likely to be developed which would further accelerate the generation and identification of useful variation for carrot quality improvement.
Bradeen, J.M. and Simon, P.W. (1998) Conversion of an AFLP fragment linked to the carrot Y2 locus to a simple, codominant, PCR-based marker form. Theor. Appl. Genet. 97: 960-967.
Just, B.J., Santos, C.A. F., Fonseca, M.E.N., Boiteux, L.S., Oloizia, B.B. and Simon, P.W. (2007) Carotenoid biosynthesis structural genes in carrot (Daucus carota): isolation, sequence-characterization, single nucleotide polymorphism (SNP) markers and genome mapping. Theor. Appl. Genet. 114: 693-704.
Just, B.J., Santos, C.A. F., Yandell, B.S. and Simon, P.W. (2009) Major QTL for carrot color are positionally associated with carotenoid biosynthetic genes and interact epistatically in a domesticated x wild carrot cross. Theor. Appl. Genet. 119: 1155-1169.
Santos, C.A.F. and Simon, P.W. (2002) QTL analyses reveal clustered loci for accumulation of major provitamin A carotenes and lycopene in carrot roots. Mol. Genet. Genomics 268: 122-129.
Simon, P.W. (1996) Inheritance and expression of purple and yellow storage root colour in carrot. Journal of Heredity 87(1): 63-66.
Vivek, B.S. and Simon, P.W. (1999) Linkage relationships among molecular markers and storage root traits of carrot (Daucus carota L. sativus). Theor. Appl. Genet. 99: 58-64.
Yau, Y.Y. and Simon, P.W. (2003) A 2.5-kb insert eliminates acid soluble invertase isozyme II transcript in carrot (Daucus carota L.) roots, causing high sucrose accumulation. Plant Mol. Biol. 53: 151-162.
Yau, Y.Y., Santos, K. and Simon, P. (2005) Molecular tagging and selection for sugar type in carrot roots using co-dominant, PCR-based markers. Molecular Breeding 16: 1-10.
About Author / Additional Info:
Working as a Senior Scientist at ICAR-Indian Agricultural Research Institute, Pusa, New Delhi