<div style="text-align:justify;"> <em>Citrus tristeza virus</em> (CTV) is an important citrus pathogen causing considerable economic loss to citrus production. Knowledge on genetic evolutio...<div style="text-align:justify;"> <em>Citrus tristeza virus</em> (CTV) is an important citrus pathogen causing considerable economic loss to citrus production. Knowledge on genetic evolutionary of the CTV population in China remains limited. In this study, 1439 samples were collected from nine citrus-producing areas of China. The coat protein (CP) genes of CTV were amplified by RT-PCR, and sequenced to analyze the genetic evolution. Analysis of the base composition showed an AU preference pattern, with the GC content was lower than AU content. Nine CTV populations were clustered into one clade in neighbor-joining (NJ) tree, indicative of a close phylogenetic relationship among the populations in China. Analysis of molecular variation (AMOVA) revealed that 77.72% genetic variations of CTV populations were observed among populations, with an <em>F</em><sub>ST</sub> value of 0.223. The values of <em>d<sub>N</sub>/d<sub>S</sub></em> and neutrality test of <em>CP</em> gene were ranged from 0.016 to 0.082 and -1.377 to 1.456, respectively, the results suggesting that all of nine CTV populations were relatively constantly maintained under purifying selection. Our study demonstrated the genetic characteristics and molecular evolution relationship of CTV populations in China, and provided a theoretical basis for scientific control of CTV. </div>展开更多
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 originati...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.展开更多
CWMV-CP1 target gene and bar selection gene were co-transferred into commercial wheat variety of Yangmai158 by particle bombardment. In total, 145 resistant plants to 3 -5 mg L-1 Bialaphos were obtained, 21 plants wer...CWMV-CP1 target gene and bar selection gene were co-transferred into commercial wheat variety of Yangmai158 by particle bombardment. In total, 145 resistant plants to 3 -5 mg L-1 Bialaphos were obtained, 21 plants were identified to be positive in T0 generation by PCR-Southern test, and the transformation frequency had 0.99%. T1 plants were further tested by PCR and Southern hybridization. Results demonstrated that the alien resistance gene had been integrated into the wheat genome. The segregation ratio of CP1+ to CP1- in T1 generation was 1.0 to 1. 3, and didn't agree with Mendelian rule. RT-PCR result from T2 plants showed that the alien gene CWMV-CP1 had stable expression in wheat genetic background.展开更多
Complementary DNAs to turnip mosaic virus (a radish Raphanus stativus isolate) genomicRNA were synthesized using oligo (dT) as primer and cloned into the vector λ-ZAP Ⅱ. Afterhybridization with a single-stranded cDN...Complementary DNAs to turnip mosaic virus (a radish Raphanus stativus isolate) genomicRNA were synthesized using oligo (dT) as primer and cloned into the vector λ-ZAP Ⅱ. Afterhybridization with a single-stranded cDNA probe and the sequencing of inserted DNA,positive clones with poly--A tails were obtained. One clone containing 1429-base pair insertwas sequenced. The coat protein gene was identified based on the molecular weight of theTuMV coat protein and the consensus sequences of the polyprotein processing sites ofpotyviruses. The 5’ end of the coat protein gene was modified by PCR to introduce aninitiation codon, ATG, and two restriction enzyme sites. The gene was then manipulatedinto a binary vector pBIN437 which was derived from pBI121, and the plant expressionvector is being used to transform Brassica napus.展开更多
To understand the effect of rare codon on the silencing ratio of foreign gene, some preferred codon in potato virus X (PVX) coat protein gene (cp) were substituted with synonymous rare codons. The modified PVX coat pr...To understand the effect of rare codon on the silencing ratio of foreign gene, some preferred codon in potato virus X (PVX) coat protein gene (cp) were substituted with synonymous rare codons. The modified PVX coat protein gene (cpm) and wild-type cp gene (cpw) were inserted into binary vector under the control of CaMV35S promoter, and these two plant expression constructs were transferred into tobacco (Nicotiana tabacum cv. Xanthi) genomes via Agrobacterium mediated method and transgenic plants were generated. Northern blot analysis of RNA isolated from these plants showed that the silencing ratio of cpm gene in transgenic tobaccos was higher than that of cpw (35% and 6.25% respectively). Run on results indicate that the silencing of cp gene happened at post-transcriptional level. The resistance of transgenic tobaccos carrying cpm genes to PVX is increased compared with that of transformants carrying cpw genes. These results suggest that the resistance of transgenic tobacco to PVX can be enhanced by codon replacement.展开更多
Ⅰ. INTRODUCTIONRice stripe disease is one of the main diseases to rice. Its pathogeny is rice stripe virus (called RSV for short). RSV can damage 37 species of gramineal crops, such as rice, wheat and corn, and decre...Ⅰ. INTRODUCTIONRice stripe disease is one of the main diseases to rice. Its pathogeny is rice stripe virus (called RSV for short). RSV can damage 37 species of gramineal crops, such as rice, wheat and corn, and decrease the yield seriously. In 1975 Koganezawa first reported that RSV possessed the branched filamentous particle. Then a series of papers was展开更多
Yam mosaic virus (YMV), a Potyvirus, is a highly destructive pathogen of yam accounting for yield losses up to 40%. Apart from causing significant reduction in tuber size and quality, it restricts international exchan...Yam mosaic virus (YMV), a Potyvirus, is a highly destructive pathogen of yam accounting for yield losses up to 40%. Apart from causing significant reduction in tuber size and quality, it restricts international exchange of germplasms. It thus becomes crucial to get resistant or at least virus-free planting materials for farmers. This study was aimed at inducing resistance to YMV in tobacco by RNA silencing. An RNAi construct containing 161 bp fragment of <span style="font-family:Verdana;">YMV-coat protein (CP) </span><span style="font-family:Verdana;">gene was developed and used to produce transgenic tobacco lines expressing </span><span style="font-family:Verdana;">YMV-coat protein (CP)</span><span style="font-family:Verdana;"> derived </span><span style="font-family:Verdana;">double stranded RNA (dsRNA)</span><span style="font-family:""><span style="font-family:Verdana;"> via </span><i><span style="font-family:Verdana;">Agrobacterium</span></i><span style="font-family:Verdana;">-mediated transformation. Of the eight T</span><sub><span style="font-family:Verdana;">1</span></sub><span style="font-family:Verdana;"> transgenic lines inoculated with YMV, six (L1, L2, L3, L5, L7 and L8) showed immunity to YMV as no symptoms were detected, whereas two (L4 and L10) exhibited high resistance with mild symptoms limited to inoculation portions. No virus could be detected in uninoculated new leaves of the transgenic lines after RT-PCR and qPCR analyses of </span></span><span style="font-family:Verdana;">YMV-coat protein (CP)</span><span style="font-family:Verdana;">. </span><span style="font-family:Verdana;">The presence of small interfering RNAs in transgenic</span><span style="font-family:MinionPro-Capt;"> </span><span style="font-family:Verdana;">lines after virus challenge indicates</span><span style="font-family:Verdana;"> that the resistance was acquired through RNA silencing.</span>展开更多
In recent years, the rapid development of techniques in plant genetic engineering has provided a new way for breeding virus-resistance plants. Powell et al. constructed the cDNA gene encoding the coat protein(CP)of to...In recent years, the rapid development of techniques in plant genetic engineering has provided a new way for breeding virus-resistance plants. Powell et al. constructed the cDNA gene encoding the coat protein(CP)of tobacco mosaic virus(TMV)under the control of a plant constitutive promoter and transferred it into tobacco cells. Transgenic tobacco plants expressing TMV coat protein gene showed the resistance or delaying of disease symptom development. Subsequently, the same method was successfully used for some other viruses such as cucumber mosaic virus(CMV), alfalfa mosaic virus(ALMV), potato virus X(PVX)and potato virus Y(PVY).展开更多
文摘<div style="text-align:justify;"> <em>Citrus tristeza virus</em> (CTV) is an important citrus pathogen causing considerable economic loss to citrus production. Knowledge on genetic evolutionary of the CTV population in China remains limited. In this study, 1439 samples were collected from nine citrus-producing areas of China. The coat protein (CP) genes of CTV were amplified by RT-PCR, and sequenced to analyze the genetic evolution. Analysis of the base composition showed an AU preference pattern, with the GC content was lower than AU content. Nine CTV populations were clustered into one clade in neighbor-joining (NJ) tree, indicative of a close phylogenetic relationship among the populations in China. Analysis of molecular variation (AMOVA) revealed that 77.72% genetic variations of CTV populations were observed among populations, with an <em>F</em><sub>ST</sub> value of 0.223. The values of <em>d<sub>N</sub>/d<sub>S</sub></em> and neutrality test of <em>CP</em> gene were ranged from 0.016 to 0.082 and -1.377 to 1.456, respectively, the results suggesting that all of nine CTV populations were relatively constantly maintained under purifying selection. Our study demonstrated the genetic characteristics and molecular evolution relationship of CTV populations in China, and provided a theoretical basis for scientific control of CTV. </div>
文摘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.
基金Thig study was supported by Europe Confederation International Cooperation with Developing Countries(ECINCO)Program(IC18-CT96-0049)Chinese“863”Program(2001-AA212111).
文摘CWMV-CP1 target gene and bar selection gene were co-transferred into commercial wheat variety of Yangmai158 by particle bombardment. In total, 145 resistant plants to 3 -5 mg L-1 Bialaphos were obtained, 21 plants were identified to be positive in T0 generation by PCR-Southern test, and the transformation frequency had 0.99%. T1 plants were further tested by PCR and Southern hybridization. Results demonstrated that the alien resistance gene had been integrated into the wheat genome. The segregation ratio of CP1+ to CP1- in T1 generation was 1.0 to 1. 3, and didn't agree with Mendelian rule. RT-PCR result from T2 plants showed that the alien gene CWMV-CP1 had stable expression in wheat genetic background.
文摘Complementary DNAs to turnip mosaic virus (a radish Raphanus stativus isolate) genomicRNA were synthesized using oligo (dT) as primer and cloned into the vector λ-ZAP Ⅱ. Afterhybridization with a single-stranded cDNA probe and the sequencing of inserted DNA,positive clones with poly--A tails were obtained. One clone containing 1429-base pair insertwas sequenced. The coat protein gene was identified based on the molecular weight of theTuMV coat protein and the consensus sequences of the polyprotein processing sites ofpotyviruses. The 5’ end of the coat protein gene was modified by PCR to introduce aninitiation codon, ATG, and two restriction enzyme sites. The gene was then manipulatedinto a binary vector pBIN437 which was derived from pBI121, and the plant expressionvector is being used to transform Brassica napus.
基金supported by the National Key Basic Research Project(Grant No.G2000016205)the National Natura1 Science Foundation of China(Grant No.39989001)
文摘To understand the effect of rare codon on the silencing ratio of foreign gene, some preferred codon in potato virus X (PVX) coat protein gene (cp) were substituted with synonymous rare codons. The modified PVX coat protein gene (cpm) and wild-type cp gene (cpw) were inserted into binary vector under the control of CaMV35S promoter, and these two plant expression constructs were transferred into tobacco (Nicotiana tabacum cv. Xanthi) genomes via Agrobacterium mediated method and transgenic plants were generated. Northern blot analysis of RNA isolated from these plants showed that the silencing ratio of cpm gene in transgenic tobaccos was higher than that of cpw (35% and 6.25% respectively). Run on results indicate that the silencing of cp gene happened at post-transcriptional level. The resistance of transgenic tobaccos carrying cpm genes to PVX is increased compared with that of transformants carrying cpw genes. These results suggest that the resistance of transgenic tobacco to PVX can be enhanced by codon replacement.
文摘Ⅰ. INTRODUCTIONRice stripe disease is one of the main diseases to rice. Its pathogeny is rice stripe virus (called RSV for short). RSV can damage 37 species of gramineal crops, such as rice, wheat and corn, and decrease the yield seriously. In 1975 Koganezawa first reported that RSV possessed the branched filamentous particle. Then a series of papers was
文摘Yam mosaic virus (YMV), a Potyvirus, is a highly destructive pathogen of yam accounting for yield losses up to 40%. Apart from causing significant reduction in tuber size and quality, it restricts international exchange of germplasms. It thus becomes crucial to get resistant or at least virus-free planting materials for farmers. This study was aimed at inducing resistance to YMV in tobacco by RNA silencing. An RNAi construct containing 161 bp fragment of <span style="font-family:Verdana;">YMV-coat protein (CP) </span><span style="font-family:Verdana;">gene was developed and used to produce transgenic tobacco lines expressing </span><span style="font-family:Verdana;">YMV-coat protein (CP)</span><span style="font-family:Verdana;"> derived </span><span style="font-family:Verdana;">double stranded RNA (dsRNA)</span><span style="font-family:""><span style="font-family:Verdana;"> via </span><i><span style="font-family:Verdana;">Agrobacterium</span></i><span style="font-family:Verdana;">-mediated transformation. Of the eight T</span><sub><span style="font-family:Verdana;">1</span></sub><span style="font-family:Verdana;"> transgenic lines inoculated with YMV, six (L1, L2, L3, L5, L7 and L8) showed immunity to YMV as no symptoms were detected, whereas two (L4 and L10) exhibited high resistance with mild symptoms limited to inoculation portions. No virus could be detected in uninoculated new leaves of the transgenic lines after RT-PCR and qPCR analyses of </span></span><span style="font-family:Verdana;">YMV-coat protein (CP)</span><span style="font-family:Verdana;">. </span><span style="font-family:Verdana;">The presence of small interfering RNAs in transgenic</span><span style="font-family:MinionPro-Capt;"> </span><span style="font-family:Verdana;">lines after virus challenge indicates</span><span style="font-family:Verdana;"> that the resistance was acquired through RNA silencing.</span>
文摘In recent years, the rapid development of techniques in plant genetic engineering has provided a new way for breeding virus-resistance plants. Powell et al. constructed the cDNA gene encoding the coat protein(CP)of tobacco mosaic virus(TMV)under the control of a plant constitutive promoter and transferred it into tobacco cells. Transgenic tobacco plants expressing TMV coat protein gene showed the resistance or delaying of disease symptom development. Subsequently, the same method was successfully used for some other viruses such as cucumber mosaic virus(CMV), alfalfa mosaic virus(ALMV), potato virus X(PVX)and potato virus Y(PVY).
