Are you Ryoi Fujiyama?

Claim your profile

Publications (5)7.06 Total impact

  • [Show abstract] [Hide abstract]
    ABSTRACT: High temperature-induced bolting of lettuce is undesirable agriculturally, making it important to find the mechanism governing the transition from vegetative to reproductive growth. FLOWERING LOCUS T (FT) genes play important roles in the induction of flowering in several plant species. To clarify floral induction in lettuce, we isolated the FT gene (LsFT) from lettuce. Sequence analysis and phylogenetic relationships of LsFT revealed considerable homology to FT genes of Arabidopsis, tomato, and other species. LsFT induced early flowering in transgenic Arabidopsis, but was not completely effective compared to AtFT. LsFT mRNA was abundant in the largest leaves under flowering-inducible conditions (higher temperatures). Gene expression was correlated with flower differentiation of the shoot apical meristem. Our results suggest that LsFT is a putative FT homolog in lettuce that regulates flower transition, similar to its homolog in Arabidopsis. This is the first information on the lettuce floral gene for elucidating regulation of the flowering transition in lettuce.
    Journal of plant physiology 03/2011; 168(13):1602-7. · 2.50 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: Lettuce big-vein disease, which is found in major lettuce production areas worldwide, is caused by Mirafiori lettuce big-vein virus (MLBVV). In order to produce marker-free transgenic lettuce resistant to MLBVV, we constructed a two T-DNA binary vector in which the first T-DNA contained a selectable marker gene npt II (neomycin phosphotransferase II) and the second T-DNA contained polyubiquitin gene promoter/terminator and inverted repeats of the viral coat protein (CP) gene. These T-DNAs were transferred via Agrobacterium tumefaciens (LBA4404)-mediated transformation into a lettuce cultivar ‘Watson’. About 40% of regenerated plants (T0 generation) on kanamycin medium showed CP gene-positive by PCR analysis. CP gene-positive plants were self-pollinated, and 124 T1 lines were analyzed for resistance to MLBVV. Twenty-one lines were selected as resistant to MLBVV, and 5 of 21 lines contained npt II-negative plants. Npt II-negative plants were self-pollinated, and 5 T2 lines were analyzed for resistance to MLBVV. All lines showed resistance to MLBVV, but one line showed npt II-positive by PCR analysis (selection error in T1 generation). The other 4 lines showed npt II-negative by both PCR and Southern blot analysis. Southern blot analysis showed that 2 of 4 lines contained one copy of the transgene per genome and that the other 2 lines contained more than one copy.
    2010 ASHS Annual Conference; 08/2010
  • [Show abstract] [Hide abstract]
    ABSTRACT: Lettuce big-vein disease is a soil-borne viral disease found in major lettuce production areas worldwide. We have developed a transgenic lettuce line with resistance to the disease by introducing the inverted repeats of the coat protein gene of Mirafiori lettuce virus (MiLV). In this study, the transgenic lettuce was evaluated for biosafety assessment by comparing its characteristics with those of a parental non-transformed cultivar ‘Kaiser’. There was no significant difference between the transgenic and the non-transgenic lettuce in pollen fertility, pollen dispersal, seed productivity, seed dispersal, dormancy, germination, growth of seedlings under low or high temperature, chromatographic patterns of leaf extracts, or effects of lettuce plants on growth of broccoli or on soil microflora. There was a significant difference in pollen size, but the difference was small. The length of cotyledon of the transgenic lettuce was a little shorter than that of ‘Kaiser,’ while there was no difference in other morphological characteristics examined. Next we detected the CP gene and the npt II gene (kanamycin resistance gene) by PCR in the transgenic lettuce of T1, T2 and T3 generations. The CP gene was detected in the transgenic lettuce of all generations. On the other hand, the npt II gene was detected in T1 plants, but not in the transgenic lettuce of T2 or T3 generation. DNA sequences flanking T-DNA insertions in the transgenic lettuce of T2 generation were determined using inverse PCR, and the result indicated that the right side of the T-DNA including the npt II gene has been truncated in the transgenic lettuce.
    2009 ASHS Annual Conference; 07/2009
  • [Show abstract] [Hide abstract]
    ABSTRACT: Lettuce big-vein disease is caused by Mirafiori lettuce virus (MiLV), which is vectored by the soil-borne fungus Olpidium brassicae. A MiLV-resistant transgenic lettuce line was developed through introducing inverted repeats of the MiLV coat protein (CP) gene. Here, a detailed characterization study of this lettuce line was conducted by comparing it with the parental, non-transformed 'Kaiser' cultivar. There were no significant differences between transgenic and non-transgenic lettuce in terms of pollen fertility, pollen dispersal, seed production, seed dispersal, dormancy, germination, growth of seedlings under low or high temperature, chromatographic patterns of leaf extracts, or effects of lettuce on the growth of broccoli or soil microflora. A significant difference in pollen size was noted, but the difference was small. The length of the cotyledons of the transgenic lettuce was shorter than that of 'Kaiser,' but there were no differences in other morphological characteristics. Agrobacterium tumefaciens used for the production of transgenic lettuce was not detected in transgenic seeds. The transgenic T(3), T(4), and T(5) generations showed higher resistance to MiLV and big-vein symptoms expression than the resistant 'Pacific' cultivar, indicating that high resistance to lettuce big-vein disease is stably inherited. PCR analysis showed that segregation of the CP gene was nearly 3:1 in the T(1) and T(2) generations, and that the transgenic T(3) generation was homozygous for the CP gene. Segregation of the neomycin phosphotransferase II (npt II) gene was about 3:1 in the T(1) generation, but the full length npt II gene was not detected in the T(2) or T(3) generation. The segregation pattern of the CP and npt II genes in the T(1) generation showed the expected 9:3:3:1 ratio. These results suggest that the fragment including the CP gene and that including the npt II gene have been integrated into two unlinked loci, and that the T(1) plant selected in our study did not have the npt II gene. DNA sequences flanking T-DNA insertions in the T(2) generation were determined using inverse PCR, and showed that the right side of the T-DNA including the npt II gene had been truncated in the transgenic lettuce.
    Transgenic Research 07/2009; 19(2):211-20. · 2.28 Impact Factor
  • Yoichi Kawazu, Ryoi Fujiyama, Yuji Noguchi
    [Show abstract] [Hide abstract]
    ABSTRACT: Mirafiori lettuce virus (MiLV), a plant RNA virus belonging to the genus Ophiovirus, is considered to be a causal agent of lettuce big-vein disease. In this study, inverted repeats of a fragment of the coat protein (CP) gene of MiLV in a binary vector pBI121 were transferred via Agrobacterium tumefaciens-mediated transformation into lettuce (Lactuca sativa L.) in order to generate MiLV-resistant lettuce. Forty T(1) lines were analyzed for resistance to MiLV by detecting MiLV in leaves, and two lines (lines 408 and 495) were selected as resistant to MiLV. Both lines were susceptible to Lettuce big-vein associated virus (LBVaV), and line 495 showed higher resistance to MiLV than line 408. Further analysis indicated that line 495 showed resistance to big-vein symptoms expression. Small interfering RNA (siRNA) molecules derived from the transgene were detected in plants of line 495. MiLV was detected in roots but not in leaves of line 495 plants after MiLV inoculation, suggesting that resistance to MiLV is less effective in roots than in leaves.
    Transgenic Research 08/2008; · 2.28 Impact Factor