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D. Gonsalves C. Gonsalves J. Carr S. Tripathi T. Matsumoto J. Suzuki S. Ferreira K. Pitz 《Tropical plant biology》2012,5(2):153-160
In 1992, papaya ringspot virus (PRSV) was discovered in the Puna district of Hawaii island where 95% of the state of Hawaii’s papaya was being grown. By 1998 production in Puna had decreased 50% from 1992 levels. A PRSV-resistant transgenic papaya ‘Rainbow’ containing the coat protein gene of PRSV was released commercially in Hawaii in 1998, and saved the papaya industry from further devastation. In the ensuing years since the release of the transgenic papaya, a number of farmers grew hermaphrodite nontransgenic ‘Kapoho’ papaya in close proximity to plantings of hermaphrodite transgenic ‘Rainbow’ papaya. These plantings provided a unique opportunity to assay for transgenic-pollen drift under commercial conditions. Between 2004 and 2010, assays for the GUS (beta-glucuronidase) transgene in embryos were done to study transgenic-pollen drift in commercial ‘Kapoho’ plantings and in replicated field plots. Very low pollen drift (0.8%) was detected in fruit of ‘Kapoho’ trees in the border row of one plantation when 90 embryos were assayed per fruit, while no pollen drift was detected in four other commercial plantings in which eight embryos were tested per fruit. Pollen drift averaged 1.3% of tested embryos in field plots where individual hermaphrodite ‘Kapoho’ trees were adjacent to two or four ‘Rainbow’ trees. In contrast, 67.4% of tested embryos were GUS positive in similarly located female ‘Kapoho’ trees. The very low transgene flow to close-by ‘Kapoho’ plantings is likely due to the fact that hermaphrodite trees are used commercially in Hawaii and that these trees are largely self-pollinated before the stigma is exposed to external pollen. 相似文献
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Transgene-specific and event-specific molecular markers for characterization of transgenic papaya lines resistant to Papaya ringspot virus 总被引:1,自引:0,他引:1
Ming-Jen Fan Shu Chen Yi-Jung Kung Ying-Huey Cheng Huey-Jiunn Bau Tien-Tsai Su Shyi-Dong Yeh 《Transgenic research》2009,18(6):971-986
The commercially valuable transgenic papaya lines carrying the coat protein (CP) gene of Papaya ringspot virus (PRSV) and conferring virus resistance have been developed in Hawaii and Taiwan in the past decade. Prompt and sensitive
protocols for transgene-specific and event-specific detections are essential for traceability of these lines to fulfill regulatory
requirement in EU and some Asian countries. Here, based on polymerase chain reaction (PCR) approaches, we demonstrated different
detection protocols for characterization of PRSV CP-transgenic papaya lines. Transgene-specific products were amplified using
different specific primer pairs targeting the sequences of the promoter, the terminator, the selection marker, and the transgene,
and the region across the promoter and transgene. Moreover, after cloning and sequencing the DNA fragments amplified by adaptor
ligation-PCR, the junctions between plant genomic DNA and the T-DNA insert were elucidated. The event-specific method targeting
the flanking sequences and the transgene was developed for identification of a specific transgenic line. The PCR patterns
using primers designed from the left or the right flanking DNA sequence of the transgene insert in three selected transgenic
papaya lines were specific and reproducible. Our results also verified that PRSV CP transgene is integrated into transgenic
papaya genome in different loci. The copy number of inserted T-DNA was further confirmed by real-time PCR. The event-specific
molecular markers developed in this investigation are crucial for regulatory requirement in some countries and intellectual
protection. Also, these markers are helpful for prompt screening of a homozygote-transgenic progeny in the breeding program. 相似文献
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Molecular biological characterization, fruit characters, and nutrients were analyzed for T4 generation of transgenic papaya.
All transgenic papaya plants with the mutated replicase (RP) gene from papaya ringspot virus (PRSV) showed high resistance
or immunity against PRSV in the field. The RP transgene can be steadily inherited to, and expressed at RNA level, the progenies.
The growth characteristics of transgenic papaya were much better than non-transgenic papaya in the field. The non-transgenic
papaya seedlings began to show typical symptoms caused by PRSV after being inoculated with PRSV. They died quickly and never
grew to produce fruit. The adult trees developed yellow leaves and produced smaller fruits and were doomed to a slow death
after some time, while most of transgenic papaya plants (about 91.8%) did not show any symptoms caused by PRSV, and produced
more, bigger, and high quality fruits. Compared with non-transgenic plants, the fresh fruit length of T4 generation of transgenic
papaya increased 2.6%–5%, and the diameter decreased 0.6%–1.5%. The flesh thickness of fresh fruit increased 12%–15%, which
made it fitter for eating. Although the fresh fruit quality changed, there was no significant difference between transgenic
and non-transgenic papaya. The quality characteristics of dry fruit including the contents of water, lipid, N, protein, reduced
sugar, vitamin A, vitamin C, and carotene in the T4 generation of transgenic papaya were all the same as their non-transgenic
parents. This means that transgenic plants and non-transgenic plants are substantially equivalent, and the transgene has no
effect on dry fruit quality. In this study, we found that vitamin A and vitamin C in red-fleshed papaya were 1.4–1.8 and 1.78–2.07
times more than the yellow-fleshed ones, respectively, while N and protein were only 84.2%–92.1% and 82.1%–98.9% of the yellow-fleshed
ones.
Translated from Acta Ecologica Sinica, 2005, 25(12): 3301–3306 [译自: 生态学报] 相似文献
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Papaya ringspot virus (PRSV) can cause a destructive disease in papaya (Carica papaya L.). Based on observations that viral replicase (RP) gene confers resistance to virus in other plants, we designed a pair of primers and cloned the RP gene from PRSV by RT-PCR. The 3'-truncated and 5'-extended RP gene fragment was then oriented under the control of the CaMV35 S promoter and nos termination sequence in the mini Ti plasmid vector pRok to construct a plant expression vector, designated pRPTW. Papaya (C. papaya L.) cv. Tai-nong-2 embryogenic calli were transformed by Agrobacterium tumefaciens LBA4404 harboring the pRPTW vector. After selection on 100 mg/ml kanamycin, 20 putative transgenic papayas were regenerated and confirmed by PCR-Southern blot and Southern blot analyses. PRSV inoculation tests showed that the RP gene conferred resistance to PRSV in transgenic papayas and those offspring carrying the RP gene. The consistency of the presence of the RP gene and PRSV resistance indicates that replicase-mediated resistance against PRSV was attained in papaya. Possible mechanisms include RNA-mediated resistance and protein-mediated resistance, as well as others, although further studies are required. 相似文献
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Pâmela Fávero Francisco de Assis Alves Mourão Filho Liliane Cristina Libório Stipp Beatriz Madalena Januzzi Mendes 《Acta Physiologiae Plantarum》2012,34(2):471-477
The difficulty in adult tissue genetic transformation in woody species is still an obstacle to be overcome, including in most
sweet orange cultivars of the Brazilian citrus industry. This work reports that, after in vitro culture adjustments, transgenic
adventitious buds of ‘Hamlin’, ‘Pêra’, and ‘Valencia’ sweet oranges (Citrus sinensis L. Osbeck) were recovered using adult material as explant source, in genetic transformation experiments via Agrobacterium tumefaciens. The transgenic buds were identified by the GUS histochemical analysis and confirmed by PCR analysis, which indicated the
presence of an amplified fragment of 817 bp corresponding to the uidA gene sequence. The efficiencies of genetic transformation for ‘Hamlin’, ‘Pêra’, and ‘Valencia’ sweet orange cultivars were
2.5, 1.4, and 3.7%, respectively. Media supplemented with auxins and cytokinins during co-culture, and media with high concentrations
of cytokinins (3 mg L−1) during transgenic selection led to the transformation and, consequently, the regeneration of adequate number of adventitious
buds for the three cultivars. The use of sonication during the explant disinfection was not effective to reduce endophytic
contamination and reduced transformation efficiency. 相似文献
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Papaya ringspot virus (PRSV) causes severe economic losses in both cucurbits and papaya throughout the tropics and subtropics. Development of PRSV-resistant
transgenic plants faces a major hurdle in achieving resistance against geographically distinct isolates. One of the major
reasons of failing to achieve the broad-spectrum PRSV resistance is the involvement of silencing suppressor proteins of viral
origin. Here, based on sequence profile of silencing suppressor protein, HcPro, we show that PRSV-HcPro, acts as a suppressor
of RNA silencing through micro RNA binding in a dose- dependent manner. In planta expression of PRSV-HcPro affects developmental biology of plants, suggesting the interference of suppressor protein in micro
RNA-directed regulatory pathways of plants. Besides facilitating the establishment of PRSV, it showed strong positive synergism
with other heterologous viruses as well. This study provides a strategy to develop effective and stable PRSV-resistant transgenic
plants. 相似文献
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Jun-Ya Wei De-Bing Liu Xiao-Ying Li Peng Zhou 《植物学报(英文版)》2007,49(7):1062-1069
The potyvirus Papaya ringspot virus (PRSV) is an important pathogen of papaya that causes severe losses in economic crops for papaya production globally. The coat protein (CP) genes of five PRSV isolates originating from different locations in China were cloned and sequenced. The CP-coding region varied in size from 864-873 nucleotides, encoding proteins of 288-291 amino acids. The five Chinese isolates of PRSV have been characterized as papaya-infecting (PRSV-P). The CP sequences of the Chinese isolates were compared with those of previously published PRSV isolates originating from different countries at amino acid levels. A number of KE repeat boxes in the N terminus of the PRSV-CP were found in all Chinese isolates. The phylogenetic branching pattern revealed that there was certain extended grouping between geographic locations, and the Asian type probably represents the oldest population of PRSV. The information of CP genes will be useful in designing and developing durable virus resistant-PRSV transgenic papaya in China. Meanwhile broad-spectrum-virus resistant, strongly resistant-PRSV and good safe papaya lines are required. 相似文献
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Wenqi Cai Carol Gonsalves Paula Tennant Gustavo Fermin Manoel Souza Jr. Nonglak Sarindu Fuh-Jyh Jan Hai-Ying Zhu Dennis Gonsalves 《In vitro cellular & developmental biology. Plant》1999,35(1):61-69
Summary A reproducible and effective biolistic method for transforming papaya (Carica papaya L.) was developed with a transformation-regeneration system that targeted a thin layer of embryogenic tissue. The key factors
in this protocol included: 1) spreading of young somatic embryo tissue that arose directly from excised immature zygotic embryos,
followed by another spreading of the actively growing embryogenic tissue 3 d before biolistic transformation; 2) removal of
kanamycin selection from all subsequent steps after kanamycin-resistant clusters were first isolated from induction media
containing kanamycin; 3) transfer of embryos with finger-like extensions to maturation medium; and 4) transferring explants
from germination to the root development medium only after the explants had elongating root initials, had at least two green
true leaves, and were about 0.5 to 1.0 cm tall. A total of 83 transgenic papaya lines expressing the nontranslatable coat
protein gene of papaya ringspot virus (PRSV) were obtained from somatic embryo clusters that originated from 63 immature zygotic
embryos. The transformation efficiency was very high: 100% of the bombarded plates produced transgenic plants. This also represents
an average of 55 transgenic lines per gram fresh weight, or 1.3 transgenic lines per embryo cluster that was spread. We validated
this procedure in our laboratory by visiting researchers who did four independent projects to transform seven papaya cultivars
with coat protein gene constructs of PRSV strains from four different countries. The method is described in detail and should
be useful for the routine transformation and regeneration of papaya.
Based in part on a presentation at the 1997 SIVB Congress on In Vitro Biology held in Washington, DC, June 14–18, 1997. 相似文献
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A survey of the main papaya (Carica papaya L.) production fields in Venezuela during 1997, indicated that crops were heavily affected with various virus‐like symptoms. A total of 745 samples from papaya plants showing symptoms suggestive of virus infection were collected and analysed using electron microscopy and enzyme‐linked immunosorbent assay (ELISA). Papaya ringspot virus (PRSV) and Papaya mild yellowing virus (PMYV) were the most frequently found viruses, which also occurred, in mixed infections. Rhabdovirus‐like particles were found only in samples collected in Distrito Federal (D.F). Papaya mosaic virus (papMV) and Tomato spotted wild virus (TSW V) were not detected during the survey. 相似文献
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P. K. Subudhi S. Nandi C. Casal S. S. Virmani N. Huang 《TAG. Theoretical and applied genetics. Theoretische und angewandte Genetik》1998,96(6-7):941-949
The cytoplasmic genetic male-sterile (CMS) lines developed at the International Rice Research Institute are valuable in producing
tropical rice hybrids. Efficient use of CMS lines in hybrid rice production will depend on their level of genetic diversity.
Aside from morphological characterization, molecular analysis based on DNA markers can provide information on the genetic
diversity of the germplasm. The Amplified Fragment Length Polymorphism (AFLP) technique was used to fingerprint 71 CMS lines
and four rice cultivars, ‘IR64’, ‘Azucena’, ‘IR74’, and ‘FR13A’. Eleven primer pair combinations specific to the enzymes PstI and MseI were used to generate 530 AFLP markers, 176 of which were polymorphic. Each CMS line revealed a distinct fingerprint. The
AFLP marker-based dendrogram depicted genetic variation among the CMS lines. The CMS lines developed in japonica background
grouped with ‘Azucena’, a japonica cultivar. None of the CMS lines clustered with ‘FR13A’, a flood-tolerant traditional indica
variety. ‘IR64’ was found to be distinct from the other indica CMS lines and clustered with lines developed in its background.
The grouping of CMS lines into a few groups is useful for breeders in selecting genetically diverse CMS lines for hybrid rice
production and in avoiding test crossing every CMS line empirically. This study demonstrated that AFLP is a powerful and reliable
tool in determining the genetic relationships and in producing distinct fingerprints of rice cultivars.
Received: 20 December 1996 / Accepted: 9 October 1997 相似文献
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L. Hartl S. Seefelder 《TAG. Theoretical and applied genetics. Theoretische und angewandte Genetik》1998,96(1):112-116
The amplified fragment length polymorphism (AFLP) technique was used to assay eight hop cultivars. The application of fluorescent-labelled
primers proved to be a valuable tool and substituted radiolabelling. Digestion with the enzymes EcoRI/ MseI and amplification with primers having three selective bases at the 3’end resulted in distinct banding patterns for imaging
with a fluorescent scanner. A total of 523 AFLP fragments derived from eight primer combinations were analysed. On average,
18 polymorphisms per combination were displayed. The Saazer “noble” hop cultivars ‘Saazer’, ‘Tettnanger’ and ‘Spalter’ could
not be discriminated. The lowest genetic similarity (GS) between lines was computed for the bitter hops ‘Hallertauer Magnum’
and ‘Wye Target’: GS value of 0.89. The high level of genetic similarity of the analysed hop cultivars is discussed.
Received: 11 August 1997 / Accepted: 22 August 1997 相似文献
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P. Bettini E. Cosi M. G. Pellegrini L. Turbanti G. Vendramin M. Buiatti 《TAG. Theoretical and applied genetics. Theoretische und angewandte Genetik》1998,97(4):575-583
Previous work carried out in our laboratory has shown that, in tomato, the alteration of endogenous phytohormone equilibria
through the integration of Agrobacterium tumefaciens genes for auxin and cytokinin synthesis can modify the active defense response to Fusarium oxysporum f. sp. lycopersici. The susceptible cv ‘Red River’ acquires a stable competence for active defense, particularly when the phytohormone equilibrium
is altered in favour of cytokinins. Here, we analyse the expression of genes involved in the defense response against pathogens,
i.e. pathogenesis-related (PR)-protein genes, in the susceptible ‘Red River’ and resistant ‘Davis’ cultivars transgenic for
the aforementioned genes. Fungal cell-wall components, glutathione, salicylic acid and the ethylene-forming ethephon are used
as “probes” for the induction of defense processes, including ethylene production. The data obtained show that the extracellular
PR-proteins (acidic chitinase and PR-1 protein) that were inducible in the control tissue of the resistant ‘Davis’ cultivar
and not expressed in the susceptible ‘Red River’ cultivar became constitutive in the transgenic tissues of both. On the other
hand, expression of the intracellular PR-proteins (basic chitinase and β-1,3-glucanase) was found to be constitutive in all
cases, both in the control and in the transgenic cell lines of the resistant and the susceptible tomato cultivars. Ethylene
production was higher in ‘Davis’ than in ‘Red River’, and significantly increased in the transgenic cell lines, particularly
when cytokinin synthesis was altered.
Received: 25 February 1998 / Accepted: 7 April 1998 相似文献
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Four experiments were conducted to determine the effect of tradewinds in Guam, USA, on growth and gas exchange of three papaya(Carica papaya L.) cultivars. Known You 1, Sunrise,and Tainung 2 papaya seedlings at two differentstages of development were exposed to 0, 36 or 100% ambientwind. Wind exposure reduced stem height and leaf or stem dryweight in most cases, but had little influence on root growth.Net CO2assimilation (ACO2) at midday was lower for seedlingsexposed to wind than for those protected from wind. Dark respirationof exposed seedlings increased as much as 120% above that ofthe protected seedlings during the night. Wind exposure decreasedwhole plant evapotranspiration by up to 36% during the photoperiod,but increased evapotranspiration by 5887% during thenocturnal period. Responses to wind exposure were similar amongcultivars, except that growth of Tainung 2 seedlingswas less affected by wind than that of the other cultivars.Seedlings that were exposed to the various wind treatments fromgermination onwards were less influenced by wind exposure thanwere seedlings that were grown in a protected nursery beforebeing exposed to the various wind treatments. These data indicatethat: (1) ambient trade winds in Guam are strong enough to decreasethe growth of papaya seedlings; (2) plant age influences theresponse; (3) stem and leaf growth are more influenced thanroot growth; and (4) decreasedACO2 and increased dark respirationmay be partly responsible for growth reduction. Copyright 2001Annals of Botany Company Carica papaya, gas exchange, wind 相似文献
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Experiments were conducted to quantify parasitism of Colorado potato beetle,Leptinotarsa decemlineata (Say), by the egg parasitoid,Edovum puttleri Grissell, on 3 different cultivars of eggplant,Solanum melongena L. Levels of parasitism were higher (P<0.05) on ‘Black Pride’ than on other cultivars. The percentage of egg masses that
were parasitized was 1.2-fold higher (P<0.05) on ‘Black Pride’ than on ‘Harris Special’ and ‘White’. The number of eggs per
mass that were parasitized was 1.3- and 1.4- fold greater (P<0.05) on ‘Black Pride’ than on ‘Harris Special’ and ‘White’,
respectively. The percentage of eggs that were parasitized per mass and percentage of emerged adult parasitoids did not differ
(P>0.05) among cultivars; between 2.1- to 2.6- fold more females than males emerged from eggs on all cultivars during the
growing season.Edovum puttleri suppressed the 2nd generation ofL. decemlineata on ‘Black Pride’ and ‘Harris Special’, but did not suppress populations on ‘White’.
相似文献
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N. Friesen R. Fritsch K. Bachmann 《TAG. Theoretical and applied genetics. Theoretische und angewandte Genetik》1997,95(8):1229-1238
Random amplified polymorphic DNA (RAPD) and genomic in situ hybridization (GISH) methods have been used to verify the hybridogenic
origin and to identify the parental species of some ornamental cultivars in the subgenus Melanocrommyum of the genus Allium. The cultivars had been selected from seed obtained after uncontrolled pollination in breeders’ fields. The combination of
GISH analysis with RAPD markers is very suitable for testing the hybridogenic origin of plants and to ascertain the parental
species of the hybrids in such cases. As suspected, A. macleanii and A. cristophii are the parental species of ‘Globemaster’. The parental species of cultivar ‘Globus’ are A. karataviense and A. stipitatum, and not A. cristophii and A. giganteum as has been assumed on morphological grounds. Cultivars ‘Lucy Ball’ and ‘Gladiator’ are of hybrid origin, though only one
of the parental species, A. hollandicum, could be confirmed. The cultivars ‘Purple Sensation’, ‘Mount Everest’, ‘White Giant’, ‘Michael H. Hoog’ and ‘Mars’ are not
hybrids since neither GISH nor RAPD suggest the presence of a second genome. ‘Purple Sensation’ belongs to A. hollandicum, ‘Mount Everest’, ‘White Giant’ and ‘Mars’ to A. stipitatum,‘Michael H. Hoog’ to A. rosenorum.
Received: 3 July 1997 / Accepted: 9 October 1997 相似文献
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This study analyzed genetic differences of 19 cultivars selected from somaclonal variants of Syngonium podophyllum Schott along with their parents as well as seven additional Syngonium species and six other aroids using amplified fragment length polymorphism (AFLP) markers generated by 12 primer sets. Among the 19 somaclonal cultivars, ‘Pink Allusion’ was selected from ‘White Butterfly’. Tissue culture of ‘Pink Allusion’ through organogenesis resulted in the development of 13 additional cultivars. Self-pollination of ‘Pink Allusion’ obtained a cultivar, ‘Regina Red Allusion’, and tissue culture propagation of ‘Regina Red Allusion’ led to the release of five other cultivars. The 12 primer sets generated a total of 1,583 scorable fragments from all accessions, of which 1,284 were polymorphic (81.9%). The percentages of polymorphic fragments within ‘White Butterfly’ and ‘Regina Red Allusion’ groups, however, were only 1.2% and 0.4%, respectively. Jaccard's similarity coefficients among somaclonal cultivars derived from ‘White Butterfly’ and ‘Regina Red Allusion’, on average, were 0.98 and 0.99, respectively. Seven out of the 15 cultivars from the ‘White Butterfly’ group and three out of six from the ‘Regina Red Allusion’ group were clearly distinguished by AFLP analysis as unique fragments were associated with respective cultivars. The unsuccessful attempt to distinguish the remaining eight cultivars from the ‘White Butterfly’ group and three from the ‘Regina Red Allusion’ group was not attributed to experimental errors or the number of primer sets used; rather it is hypothesized to be caused by DNA methylation and/or some rare mutations. This study also calls for increased genetic diversity of cultivated Syngonium as they are largely derived from somaclonal variants. 相似文献