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1.
In 2008 and 2009 seasons, a sudden increase in Potato virus Y (PVY) incidence was recorded in foundation seed potatoes in Hokkaido, northern Japan. This increase was obvious during the field inspection and the postharvest indexing. Molecular typing revealed that besides the previously reported strains of PVYO and PVYNA‐N, the most common strain identified was the recombinant PVYNTN, with three characteristic recombinant junctions at the HC‐Pro, VPg and CP regions. No potato tuber necrotic ringspot disease (PTNRD) was observed in foundation seed potatoes in correlation with the presence of PVYNTN. Moreover, an isolate with a typical PVYNTN recombinant genome, namely Eu‐12Jp, did not induce PTNRD in 62 Japanese potato cultivars tested in both primarily and secondarily infected plants. Two cultivars carrying the extreme resistance gene Rychc were resistant to the infection with Eu‐12Jp, which presents potential sources of resistance to PVYNTN. Eu‐12Jp induced systemic mottle in potato cultivars Desiree and King Edward carrying resistance genes Ny and Nc, respectively, but induced a hypersensitive reaction in potato cultivar Maris Bard, with the Nz hypothetical resistance gene typical of the PVYZ strain group. Therefore, based on the genome structure and the reaction of the potato N resistance genes, Eu‐12Jp should be classified as PVYZ‐NTN, as described for isolates from Idaho, USA recently. This is the first report of PVYZ‐NTN in Japan and the sudden and increased occurrence of PVYNTN/PVYZ‐NTN represents a potential risk of PTNRD developing and increases the significance of PVY in Japan.  相似文献   

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4.
The gene coding for potato virus X (PVX) coat protein (CP) was expressed in transgenic potato plants obtained byAgrobacterium tumefaciens transformation. One hundred independent clones were analyzed in challenge experiments for resistance to PVX infection under greenhouse conditions as a preliminary test. From this test, 16 clones with the best resistance results were selected for a small-scale field trial. Clones 54, 60, 73 and 91 demonstrated the best values of resistance to PVX in the field. Statistical analysis of the field trial showed significant differences between means of optical density obtained in ELISA from transgenic clones and non-transformed plants (P<0.05). There was correspondence between resistance to virus infection and expression of the CP gene of PVX virus in the analyzed clones. http://www.phytoparasitica.org posting Jan. 21, 2002. Corresponding author [e-mail: vivian.doreste@cigb.edu.cu].  相似文献   

5.
Resistance tests were made on seedlings of transformed lines of Nicotiana benthamiana which contain a transgene encoding the coat protein (CP) gene of a Scottish isolate of potato mop-top virus (PMTV). This transgene has been reported to confer strong resistance to the PMTV isolate from which the transgene sequence was derived and also to a second Scottish isolate. Plants of lines of the transgenic N. benthamiana were as resistant to two Swedish and two Danish PMTV isolates as to a Scottish isolate, and of five lines tested, greater than 93.5% of transgenic plants were immune. The coat protein gene sequences of these four Scandinavian isolates were very similar to those of the two Scottish isolates. The greatest divergence between the isolates was three amino acid changes and there was less than 2% change in CP gene nucleotide sequence. It is concluded that the PMTV CP transgene used in these experiments could confer resistance against isolates from different geographical areas because it is becoming apparent that the CP genes of PMTV isolates are highly conserved.  相似文献   

6.
Transgenic tomato plants containing the coat protein (CP) gene of Cucumber mosaic virus (CMV) of subgroup IB were developed through Agrobacterium-mediated transformations. The progenies of transgenic plants showed the presence of transgene, its expression and translation of 26 KDa CP. The T1 and T2 generation plants were evaluated for resistance against challenge inoculations by a homologous strain of CMV. Visual observations of challenged transgenic plants categorized them into resistant, tolerant and susceptible as compared with untransformed control plants. Out of 33 plants of the T1 generation, 36.3% showed resistance and remained symptomless throughout their life, 48.4% showed tolerance which developed delayed symptoms of mild mosaic, and 15.1% showed susceptibility to CMV which developed severe systemic mosaic and leaf distortion symptoms after 30?days of virus challenge. Out of 120 plants of the T2 generation, 60% showed resistance, 26.6% were tolerant and only 13.3% were found susceptible to challenge inoculations of CMV. Resistant transgenic plants also showed less CP accumulation in systemic upper leaves as compared with challenged untransformed plants. In this study, CP of a CMV subgroup IB strain has demonstrated a significant level of resistance in transgenic tomato plants against the CMV strain. The strategy may be applied for better quality and productivity of tomato crops.  相似文献   

7.
Two Potato virus Y (PVY) isolates collected in Brazil, PVY‐AGA and PVY‐MON, were identified as recombinants between two parent genomes, PVYNTN and PVY‐NE‐11, with a novel type of genomic pattern. The new recombinants had an ordinary PVYNTN genome structure for approximately 6·7‐kb from the 5′‐end of the genome whereas the 3′‐terminal 3·0‐kb segment had two fragments of NE‐11‐like sequence separated by another small PVYNTN‐like fragment. PVY strains are defined based on the hypersensitive resistance (HR) response in potato indicators. Both PVY‐AGA and PVY‐MON isolates did not induce the HR in potato cultivars carrying Ny, Nc, or (putative) Nz genes and thus were able to overcome all known resistance genes to PVY. Only one of the two isolates, PVY‐AGA, induced a vein necrosis reaction in tobacco. The biological responses of the potato indicators and tobacco defined PVY‐MON as an isolate of the PVYE strain. To distinguish PVY‐AGA and PVY‐MON from other PVYNTN isolates, an RT‐PCR test was developed utilizing new specific primers from the capsid protein gene area and producing a characteristic 955‐bp band. Serological profiling of these PVY isolates with three monoclonal antibodies revealed an unusual reactivity, where one of the two commercial PVYN‐specific monoclonal antibodies did not recognize PVY‐AGA. The ability of these new PVY recombinants to overcome resistance genes in potato producing mild or no symptoms, combined with the lack of serological reactivity towards at least one PVYN‐specific antibody may present a significant threat posed by these isolates to seed potato production areas.  相似文献   

8.
An Austrian isolate of potato virus YNTN, the causal agent of potato tuber necrotic ringspot disease (PTNRD), was serologically compared with seven Dutch PVYN isolates. Using polyclonal and monoclonal antibodies, it was found indistinguishable from PVYN. Determination of the nucleotide sequence of the coat protein cistron and comparison of the deduced amino acid sequence with coat protein sequences of other potyviruses revealed a high level of homology with PVYN coat protein sequences. This confirmed the close taxonomic relationship of PVYNTN with the PVYN subgroup of potato virus Y. PVYNTN is able to overcome all resistance genes known so far in commercial potato cultivars. Remarkably, transgenic PVY-protected tobacco plants are also resistant to PVYNTN infection upon mechanical and aphid-mediated inoculation. These experiments indicate that genetically engineered resistance offers great potential in protection of potato to new aggressive strains of PVYN.  相似文献   

9.
The effect of cultivation temperatures on the resistance reaction to three Potato virus Y strains (PVYO, PVYN and PVYNTN) in potato cultivars carrying Rychc was examined. When potato plants carrying Rychc were cultivated at 22 °C, a few small necrotic spots developed on inoculated leaves by 5 days after mechanical inoculation (dpi), and systemic infection of a few symptomless plants was confirmed at 28 dpi by IC‐RT‐PCR. At 28 °C, distinct necrotic spots developed on inoculated leaves by 5 dpi, and systemic symptoms occasionally appeared at 28 dpi. Thus, high temperature weakens Rychc‐conferred resistance. However, the incidence of systemic infection and the titre of virus in resistant cultivars at 28 °C were lower than in a susceptible cultivar. In graft inoculation under high summer temperatures, some plants developed necrosis on the leaves and stem, but PVY was barely detected by RT‐PCR in leaves on potato carrying Rychc. When seedlings from progeny tubers of plants that were inoculated with PVY and grown in a greenhouse at >30 °C in the daytime were examined by ELISA and IC‐RT‐PCR, PVY was not detected in cultivars carrying Rychc. These results show that Rychc confers an extreme resistance to PVY strains occurring in Japan.  相似文献   

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12.
Tissue cultures of GAF30/Park oats were biolistically co-transformed with constructs containing the coat protein (CP) genes of the P-PAV, MAV-PS1 or NY-RPV isolates of barley yellow dwarf virus (BYDV), together with a construct containing the bar gene for herbicide resistance and the uidA reporter gene. Transformed, herbicide-resistant tissue cultures were screened by PCR for the presence of the CP genes. Fertile regenerated plants were recovered from some CP-transformed tissue cultures. T1 progeny of these plants were screened for resistance to the BYDV isolate corresponding to the introduced gene by inoculation with viruliferous aphids followed by ELISA tests. Variation in ELISA values for GAF30/Park control plants made interpretation of the data difficult, but oat plants resistant to each of the three isolates of BYDV (ELISA values less than 0.3; virus titers equivalent to less than 25% of infected controls) were identified in T1 generations. Further testing of MAV-PS1 CP-transformed lines to the T2 generation, NY-RPV CP-transformed lines to the T3 generation and P-PAV CP-transformed lines to the T4 generation identified further resistant plants. Similarly, immature embryos and calli of the barley cultivar Golden Promise were biolistically bombarded with constructs containing the CP gene of the P-PAV isolate of BYDV and the bar and uidA reporter genes, lines of self-fertile P-PAV CP-transformed barley plants were developed, and T1plants were screened for resistance to P-PAV. Eight plants from six lines showed moderate to high levels of resistance to P-PAV that correlated with the presence of the CP gene. Plants giving low ELISA values were also found in other lines, even though the CP gene was not detected in these plants. Some T2 plants derived from resistant parents that contained the CP gene were themselves highly resistant.  相似文献   

13.
hpRNA的茎环比例对RNA介导的病毒抗性产生的影响   总被引：1，自引：0，他引：1  
 以马铃薯Y病毒坏死株系(PVYN)的外壳蛋白(coat protein,CP)基因3'端50bp片段为hpRNA的茎,以pUC19不同长度的序列为hpRNA的环,构建了茎环比例分别为4:1、2:1、1:1、1:2、1:4和1:8的植物表达载体。利用农杆菌介导法转化烟草品种NC89,获得了多种转基因植株。室内抗病性检测发现:不同茎环比例的hpRNA介导的病毒抗性效率不同;茎环比例为4:1、2:1和1:1时效率较高,抗性植株的比例达60%左右;随着环长度的逐渐增加,抗性植株的比例逐渐降低;当茎环比例为1:8时,抗性植株的比例仅为9.52%。Southern blot分析结果表明:外源基因已整合于烟草的基因组中,且转基因植株的抗病性与转基因的拷贝数之间无明显的相关性。Northern blot分析结果表明:目的片段转录产物的积累量与植株的抗病性呈负相关,证明所获得的抗病性是RNA介导的。  相似文献   

14.
The influence of co‐infection on concentration and accumulation of genetically different isolates of Potato virus Y (PVY) in potato and tobacco plants and the efficiency of transmission by Myzus persicae of PVY isolates from doubly versus singly infected plants were evaluated. The vector ability to simultaneously transmit two virus isolates was examined. Eight PVY isolates represented three strain groups: PVYO (pathotype and serotype O), PVYNW (pathotype N and serotype O), and PVYNTN (pathotype and serotype N). Different diagnostic methods, including DAS‐ELISA, multiplex RT‐PCR, aphid transmission tests and bioassays, were applied to detect the presence of PVY isolates in source and assay plants. Significant reductions in concentrations of certain PVY isolates during co‐infection with other isolates were found both in potato and tobacco plants. The observed effects were both isolate‐ and host‐dependent in form. The highest rates of virus transmission by single aphids were recorded with PVYNTN isolates, and the lowest ones with PVYO isolates. Individual aphids of M. persicae were able to simultaneously transmit two PVY isolates. The frequency of transmission was generally low, but it reached as high as 20% for one of the isolate combinations. The findings presented in the work provide proof for antagonistic within‐plant interactions between isolates of PVY, with some implications of these interactions for virus transmission by aphid vectors. Consequently, this research contributes to a better understanding of the epidemiology of the disease caused by PVY.  相似文献   

15.
Surveys were conducted of symptomatic potato plants in late season crops, from the major potato production regions in Northern Tunisia, for infection with six common potato viruses. The presence of Potato leafroll virus (PLRV), Potato virus Y (PVY), Potato virus X (PVX), Potato virus A (PVA), Potato virus S (PVS) and Potato virus M (PVM) was confirmed serologically with virus infection levels up to 5.4, 90.2, 4.3, 3.8, 7.1 and 4.8%, respectively. As PVY was prevalent in all seven surveyed regions, further biological, serological and molecular typing of 32 PVY isolates was undertaken. Only one isolate was shown to induce PVYO-type symptoms following transmission to tobacco and to react only against anti-PVYO-C antibodies. Typical vein necrosis symptoms were obtained from 31 samples, six of which reacted against both anti-PVYN and anti-PVYO-C antibodies showing they contained mixed isolates, while 25 of them reacted only with anti-PVYN antibodies. An immunocapture RT-PCR molecular test using a PVYNTN specific primer pair set in the 5’NTR/P1 genomic region and examination of recombinant points in three genomic regions (HC-Pro/P3, CI/NIa and CP/3’NTR) showed that all 25 serotype-N PVY isolates were PVYNTN variants with similar recombinations to the standard PVYNTN-H isolate. This is the first report of the occurrence of the PVYNTN variant and its high incidence in late season potatoes in Tunisia.  相似文献   

16.
Cowpea (Vigna unguiculata) is one of the most important legumes cultivated in many parts of the world. The diseases caused by Cowpea severe mosaic virus (CPSMV) and Cowpea aphid‐borne mosaic virus (CABMV) are considered among the most important constraints on yield and quality, especially in Latin America and Africa. Here, the concept of using an RNA interference construct to silence the CPSMV proteinase cofactor gene and the CABMV coat protein gene is explored, in order to generate resistant transgenic cowpea plants. Ten cowpea transgenic lines were produced, presenting a normal phenotype and transferring the transgene to the next generation. Plants were tested for resistance to both CABMV and CPSMV by mechanical co‐inoculation. Seven lines presented milder symptoms when compared to the control and three lines presented enhanced resistance to both viruses. Northern analyses were carried out to detect the transgene‐derived small interfering RNA (siRNA) in leaves and revealed no correlation between siRNA levels and virus resistance. Additionally, in the symptomless resistant lines the resistance was homozygosis‐dependent. Only homozygous plants remained uninfected while hemizygous plants presented milder symptoms.  相似文献   

17.
Transgenic tobacco plants expressing a phenylalanine ammonia-lyase cDNA (ShPAL), isolated from Stylosanthes humilis, under the control of the 35S promoter of the cauliflower mosaic virus were produced to test the effect of high level PAL expression on disease resistance. The transgenic plants showed up to eight-fold PAL activity and were slowed in growth and flowering relative to non-transgenic controls which have segregated out the transgene. The expression of the ShPAL transgene and elevated PAL levels were correlated and stably inherited. In T1 and T2 tobacco plants with increased PAL activity, lesion expansion was significantly reduced by up to 55% on stems inoculated with the Oomycete pathogen Phytophthora parasitica pv. nicotianae. Lesion area was significantly reduced by up to 50% on leaves inoculated with the fungal pathogen Cercospora nicotianae. This study provides further evidence that PAL has a role in plant defence.  相似文献   

18.
The coat protein (CP) gene mediated transgenic resistance is found to be the best approach for protecting papaya plants against the destructive disease caused by Papaya ringspot viruses(PRSV). In order to study the variability of PRSV and the potential threat to the CP-transgenic resistance, five virus isolates were collected from transgenic plants of papaya line 16-0-1, which carry the CP gene of the typical mosaic strain of Taiwan PRSV YK, in an approved test field and fourteen from untransformed papaya plants in different areas of Taiwan. The results of biological, serological, and molecular characterization indicated that all isolates are related to PRSV YK. Among them, the isolate 5--19 from the transgenic line and the isolates CS and TD2 from untransformed papaya were able to overcome the YK CP gene-mediated resistance of papaya lines 18--2--4, 17-0-5, and 16-0-1, which provide high degrees of resistance to different geographic PRSV strains of Hawaii (HA), Mexico (MX), and Thailand (TH). These three isolates were also able to cause symptoms on untransformed papaya plants more severe than those induced by YK. In addition to the host reactions, the variability of the collected 19 isolates was also analyzed and compared with YK and other geographic strains by heteroduplex mobility assay (HMA) and sequence analyses. The results of HMA indicated that the CP genes of isolates 5--19 and TD2 are more divergent than those of other isolates when compared with YK. However, sequence analyses of the transgenic-resistance overcoming isolates 5-19, CS, and TD2 revealed that their CP coding regions and the 3 untranslated regions (UTRs) share nucleotide identities of 93.9–96.6% and 94.2–97.9% with those of YK, respectively; whereas the other geographic strains of HA, MX, and TH that could not overcome the transgenic resistance share lower nucleotide identities of 89.8–92.6% and 92.3–95.3% with those of YK, respectively. Our results indicate that the ability for overcoming the transgenic resistance is not solely correlated with higher degrees of sequence divergence from the transgene. The possible mechanism for overcoming the transgenic resistance and the potential threat of these PRSV strains to the application of the transgenic papaya lines carrying PRSV YK CP gene are discussed.  相似文献   

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The gene gdhA from Escherichia coli, that encodes a NADPH‐dependent glutamate dehydrogenase (GDH), directs a novel pathway in transgenic plants that allows an increase in ammonium assimilation. Glufosinate leads to plant death by the irreversible inhibition of glutamate synthetase (GS) leading to a disruption of subsequent GS‐related processes resulting in elevated ammonium and disruption of photorespiration. Therefore, it was speculated that the gdhA‐transformed plants may exhibit a novel mechanism of resistance to glufosinate by altered activity of the GDH‐directed pathway(s) and subsequently related processes. Studies were conducted in the greenhouse to evaluate the resistance of tobacco plants containing the gdhA gene to glufosinate. Five tobacco genotype lines were investigated including a non‐transformed control line, a positive control line and three transformed lines with levels of increasing GDH activity directed by the gdhA gene. Plants transformed with the gdhA gene expressed up to six times increased level of resistance (GR50) to glufosinate compared with the non‐transformed control, which is 100 times less resistant than plants transformed with the bar gene. The GDH activity among lines was highly correlated (r2 = 0.9903) with the level of herbicide resistance. Thus, the use of the E. coli gdhA gene in plant transformations can provide an additional mechanism for resistance to glufosinate.  相似文献   

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