To introduce downy mildew resistance from a yellow-colored resistant cultivar, ‘Santero’, into a yellow breeding line, OT803, the F1 hybrid was produced by crossing Santero and OT803. The bulb color of the F1 hybrids became light pink, suggesting involvement of complementation between the
Bulb color is one of major traits in onion (
Anthocyanin, one of major flavonoid compounds, is responsible for the red bulb color, and quercetin derivatives are predominant in yellow onions (Fossen
These five loci was assumed to be related with genes involved in anthocyanin biosynthesis pathway (Kim
In this study, a novel
A F1 population originating from the cross between a downy mildew resistant cultivar, ‘Santero’, and a breeding line (OT803) was used to identify a new
Total genomic DNAs were extracted from leaves of seedling or sprouted bulbs using a cetyl trimethy-lammonium bromide (CTAB) method (Doyle and Doyle 1987). PCR was performed in 25-μL reaction mixtures containing 0.05 μg template, 2.5 μL 1× PCR buffer, 0.2 μL forward primer (10 μM), 0.2 μL reverse primer (10 μM), 0.2 μL dNTPs (10 mM each), and 0.25 μL polymerase mix (Advantage 2 Polymerase Mix; Clontech, Palo Alto, CA, U.S.A.). The primer sequences used in this study are listed in Table 1.
PCR amplification was performed with an initial denaturation step at 94°C for 5 minutes, followed by 40 cycles of 94°C for 30 seconds, 65°C for 30 seconds, 72°C for 3 minutes, and a final 10 minutes extension at 72°C. The PCR products were visualized on a 1.5% agarose gel after ethidium bromide staining. For sequencing of PCR products, they were first purified using a QIAquick PCR Purification kit (QIAGEN, Valencia, CA, U.S.A.), and sequencing reactions were performed by a specialized company (Macrogen, Seoul, Republic of Korea). The full-length sequences of the new
Total RNAs were extracted from fresh leaf sheaths of four-leaf stage seedlings using a RNA extraction kit (RNeasy Plant Mini Kit, QIAGEN) following the manufacturer’s instructions. cDNAs were synthesized using a commercial cDNA synthesis kit (SuperScript™ III first-strand synthesis system for RT-PCR; Invitrogen, Carlsbad, CA, U.S.A.). RT-PCR amplification was performed with an initial denaturation step at 94°C for 3 minutes followed by 30 cycles of 94°C for 30 seconds, 65°C for 30 seconds, and 72°C for 3 minutes, and a final 10-minute extension at 72°C. The primer pair used in RT-PCR is listed in Table 1. The onion tubulin sequence obtained from EST sequences (TC125) from the DFCI
In order to introduce downy mildew resistance from a resistant cultivar, Santero, into a breeding line (OT803), the F1 hybrids were produced by the cross between Santero and OT803. Although the bulb color of both parental lines were yellow, the bulb color of the F1 hybrids became light pink (Fig. 1). The light pink bulb color was assumed to appear by complementation between the
The 4,830-bp full-length sequences including putative promoter regions of the
In the previous study (Kim
For identification of specific
In the previous study (Song
A novel inactive
However, the RT-PCR result showed that the transcripts of the
Although the causal genes responsible for the bulb color difference between red and yellow onions have been revealed as the genes coding for DFR and ANS enzymes (Kim
This research was supported by the Agriculture Research Center program, Golden Seed Project (Center for Horticultural Seed Development, No 213007-05-1-SBB10) and a grant from the Next-Generation BioGreen 21 Program (Plant Molecular Breeding Center No. PJ011034). The authors thank Ji-wha Hur, Jeong-Ahn Yoo, and Su-jung Kim for their dedicated technical assistance.
Primer sequences used in this study.
|Name||Sequence (5′ to 3′)||Application||Reference|
|OT803||Mixed with homozygous and heterozygous |