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ABSTRACT: Given that feral transgenic canola (Brassica napus) from spilled seeds has been found outside of farmer's fields and that B. juncea is distributed worldwide, it is possible that introgression to B. juncea from B. napus has occurred. To investigate such introgression, we characterized the persistence of B. napus C genome chromosome (C-chromosome) regions in backcross progenies by B. napus C-chromosome specific simple sequence repeat (SSR) markers. We produced backcross progenies from B. juncea and F(1) hybrid of B. juncea × B. napus to evaluate persistence of C-chromosome region, and screened 83 markers from a set of reported C-chromosome specific SSR markers. Eighty-five percent of the SSR markers were deleted in the BC(1) obtained from B. juncea × F(1) hybrid, and this BC(1) exhibited a plant type like that of B. juncea. Most markers were deleted in BC(2) and BC(3) plants, with only two markers persisting in the BC(3). These results indicate a small possibility of persistence of C-chromosome regions in our backcross progenies. Knowledge about the persistence of B. napus C-chromosome regions in backcross progenies may contribute to shed light on gene introgression.
Breeding Science 12/2012; 62(4):328-33. · 1.25 Impact Factor
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ABSTRACT: Several imported transgenic canola (Brassica napus) seeds have been spilled and have grown along roadsides around import ports. B. juncea, a relative of B. napus with which it has high interspecific crossability, is widely distributed throughout Japan. There is public concern about the harmful impacts of feral B. napus plants on biodiversity, but spontaneous hybridization between spilled B. napus and weedy B. juncea populations is hardly revealed. We evaluated the relationship between the hybridization frequency of B. juncea × B. napus and their planting distance in field experiments using the mutagenic herbicide-tolerant B. napus cv. Bn0861 as a pollen source for hybrid screening. The recipient B. juncea cv. Kikarashina was planted in an experimental field with Bn0861 planted in the center. No hybrids were detected under natural flowering conditions in 2009. However, the flowering period was artificially kept overlapping in 2010, leading to a hybridization frequency of 1.62% in the mixed planting area. The hybridization frequency decreased drastically with distance from the pollen source, and was lower under field conditions than estimated from the high crossability, implying that spontaneous hybridization between spilled B. napus and weedy B. juncea is unlikely in the natural environment.
Breeding Science 09/2012; 62(3):274-81. · 1.25 Impact Factor
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ABSTRACT: Imported genetically modified (GM) canola (Brassica napus) is approved by Japanese law. Some GM canola varieties have been found around importation sites, and there is public concern that these may have any harmful effects on related species such as reduction of wild relatives. Because B. juncea is distributed throughout Japan and is known to be high crossability with B. napus, it is assumed to be a recipient of B. napus. However, there are few reports for introgression of cross-combination in B. juncea × B. napus. To assess crossability, we artificially pollinated B. juncea with B. napus. After harvesting a large number of progeny seeds, we observed false hybrids and metaxenia of seed coats. Seed coat color was classified into four categories and false hybrids were confirmed by morphological characteristics and random amplified polymorphic DNA (RAPD) markers. Furthermore, the occurrence of false hybrids was affected by varietal differences in B. napus, whereas that of metaxenia was related to hybridity. Therefore, we suggest that metaxenia can be used as a marker for hybrid identification in B. juncea L. cv. Kikarashina × B. napus. Our results suggest that hybrid productivity in B. juncea × B. napus should not be evaluated by only seed productivity, crossability ought to be assessed the detection of true hybrids.
Breeding Science 12/2011; 61(4):358-65. · 1.25 Impact Factor
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ABSTRACT: An efficient genetic transformation method for kabocha squash (Cucurbita moschata Duch cv. Heiankogiku) was established by wounding cotyledonary node explants with aluminum borate whiskers prior to inoculation with Agrobacterium. Adventitious shoots were induced from only the proximal regions of the cotyledonary nodes and were most efficiently induced on Murashige-Skoog agar medium with 1 mg/L benzyladenine. Vortexing with 1% (w/v) aluminum borate whiskers significantly increased Agrobacterium infection efficiency in the proximal region of the explants. Transgenic plants were screened at the T(0) generation by sGFP fluorescence, genomic PCR, and Southern blot analyses. These transgenic plants grew normally and T(1) seeds were obtained. We confirmed stable integration of the transgene and its inheritance in T(1) generation plants by sGFP fluorescence and genomic PCR analyses. The average transgenic efficiency for producing kabocha squashes with our method was about 2.7%, a value sufficient for practical use.
Plant Cell Reports 03/2011; 30(8):1455-64. · 2.27 Impact Factor
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ABSTRACT: We investigated estrogen-inducible green fluorescent protein (GFP) expression patterns using an estrogen receptor fused chimeric transcription activator, XVE, in the monocotyledonous model plant rice (Oryza sativa L.). This system has been shown to be an effective chemical-inducible gene expression system in Arabidopsis and has been applied to other plants in order to investigate gene functions or produce marker-free transgenic plants. However, limited information is available on the correlation between inducer concentration and the expression level of the gene induced in monocots. Here, we produced a transgenic rice integrated estrogen-inducible GFP expression vector, pLex:GFP, and investigated dose-response and time-course patterns of GFP induction in rice calli and seedlings for the first time. With 17-β-estradiol treatment at >5 μM, GFP signals were detected in the entire surface of calli within 2 days of culture. Highest GFP signals were extended for 8 days with estradiol treatment at 25 μM. In three-leaf-stage seedlings, GFP signals in the leaves of pLex:GFP-integrated transgenic lines were weaker than those in the leaves of p35S:GFP-integrated transgenic lines. However, GFP signals in the roots of pLex:GFP- and p35S:GFP-integrated transgenic lines were similar with estradiol treatment at >10 μM. With regard to controlling appropriate gene expression, these results might provide helpful indications on estradiol treatment conditions to be used for the XVE system in rice and other monocots.
Plant Cell Reports 12/2010; 30(4):529-38. · 2.27 Impact Factor
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ABSTRACT: Prevention of transgene flow from genetically modified crops to food crops and wild relatives is of concern in agricultural biotechnology. We used genes derived from food crops to produce complete male sterility as a strategy for gene confinement as well as to reduce the food purity concerns of consumers. Anther-specific promoters (A3, A6, A9, MS2, and MS5) were isolated from Brassica oleracea and B. rapa and fused to the beta-glucuronidase (GUS) reporter gene and candidate genes for male sterility, including the cysteine proteases BoCysP1 and BoCP3, and negative regulatory components of phytohormonal responses involved in male development. These constructs were then introduced into Arabidopsis thaliana. GUS analyses revealed that A3, A6, and A9 had tapetum-specific promoter activity from the anther meiocyte stage. Male sterility was confirmed in tested constructs with protease or gibberellin insensitive (gai) genes. In particular, constructs with BoCysP1 driven by the A3 or A9 promoter most efficiently produced plants with complete male sterility. The tapetum and middle layer cells of anthers expressing BoCysP1 were swollen and excessively vacuolated when observed in transverse section. This suggests that the ectopic expression of cysteine protease in the meiocyte stage may inhibit programmed cell death. The gai gene also induced male sterility, although at a low frequency. This is the first report to show that plant cysteine proteases and gai from food crops are available as a novel tool for the development of genetically engineered male-sterile plants.
Plant Cell Reports 09/2008; 27(11):1741-54. · 2.27 Impact Factor
