Firmness is one of the most important fruit quality traits in strawberries.The postharvest shelf life of this soft fruit is highly limited by the loss of firmness,where cell wall disassembly plays an important role.Pr...Firmness is one of the most important fruit quality traits in strawberries.The postharvest shelf life of this soft fruit is highly limited by the loss of firmness,where cell wall disassembly plays an important role.Previous studies demonstrated that the polygalacturonase FaPG1 has a key role in remodelling pectins during strawberry softening.In this study,FaPG1 knockout strawberry plants have been generated using the CRISPR/Cas9 system delivered via Agrobacterium tumefaciens.Ten independent lines,cv.“Chandler”,were obtained,and all of them were successfully edited as determined by PCR amplification and T7 endonuclease assay.The targeted mutagenesis insertion and deletion rates were analyzed using targeted deep sequencing.The percentage of edited sequences varied from 47%up to almost 100%,being higher than 95%for seven of the selected lines.Phenotypic analyses showed that 7 out of the eight lines analyzed produced fruits significantly firmer than the control,ranging from 33 to 70%increase in firmness.There was a positive relationship between the degree of FaPG1 editing and the rise in fruit firmness.Minor changes were observed in other fruit quality traits,such as colour,soluble solids,titratable acidity or anthocyanin content.Edited fruits showed a reduced softening rate during postharvest,displayed a reduced transpirational water loss,and were less damaged by Botrytis cinerea inoculation.The analysis of four potential off-target sites revealed no mutation events.In conclusion,editing the FaPG1 gene using the CRISPR/Cas9 system is an efficient method for improving strawberry fruit firmness and shelf life.展开更多
In this study, 1 500 bp PG gene promoter was amplified from leaves of tomato cultivar ＇ Zhongshu No. 4＇. The bioinformatic analysis of PG promoter sequences was conducted. PG promoter elements were predicted and ana...In this study, 1 500 bp PG gene promoter was amplified from leaves of tomato cultivar ＇ Zhongshu No. 4＇. The bioinformatic analysis of PG promoter sequences was conducted. PG promoter elements were predicted and analyzed by PLANTCARE and PLACE. The results showed that tomato PG promoter contained multiple c/s-acting regulatory elements such as typical basic elements TATA-Box and CAAT-Box, light responsive elements 3-AF1 binding site, ATl-motif, ATCT- motif, Box 4, Box I, GA-motif, GTl-motif, Spl and MRE, heat stress-responsive element HSE, ethylene-responsive element ERE, meristem-specifie regulatory element CCGTCC-box, endosperm expression-related regulatory elements GCN4_motif and Skn-l_motif, defense and stress responsiveness element TC-rich repeats, and circadian control-related element circadian, indicating that the expression of tomato PG gene is related to light, temperature, hormone, stress and other factors. This study laid the foundation for subsequent research about regulation of PG gene expression in plants.展开更多
目的:探讨腹腔镜胃癌D2根治术患者经协同护理及患者自评-主观全面评定(patient-generated subjective global assessment,PG-SGA)个体化营养干预的效果。方法:选择蚌埠医科大学第一附属医院2020年8月—2021年11月收治的107例腹腔镜胃癌D...目的:探讨腹腔镜胃癌D2根治术患者经协同护理及患者自评-主观全面评定(patient-generated subjective global assessment,PG-SGA)个体化营养干预的效果。方法:选择蚌埠医科大学第一附属医院2020年8月—2021年11月收治的107例腹腔镜胃癌D2根治术患者,随机分为参照组(n=54,行常规护理)和试验组(n=53,行协同护理+基于PG-SGA个体化营养干预)。对比两组患者的心理状态及生活质量。结果:试验组干预后汉密尔顿焦虑量表(Hamilton anxiety scale,HAMA)、汉密尔顿抑郁量表(Hamilton depression scale,HAMD)评分均较参照组低,健康调查简表(the MOS item short form health survey,SF-36)各维度评分均较参照组高,差异均有统计学意义(P<0.05)。结论:协同护理联合基于PG-SGA个体化营养干预应用于腹腔镜胃癌D2根治术患者,能够有效改善患者的心理状态,提升其生活质量。展开更多
Polygalacturonase (PG,EC3.2.1.15) is the key cell wall hydrolase in fruit ripening. The identification and characterization of a full length cDNA (pMT18) encoding for PG from Feicheng peach (Prunus persica (L.) Bat...Polygalacturonase (PG,EC3.2.1.15) is the key cell wall hydrolase in fruit ripening. The identification and characterization of a full length cDNA (pMT18) encoding for PG from Feicheng peach (Prunus persica (L.) Batsch cv. Feicheng) is described. The pMT18 clone is 1188 bp in length, with an open reading frame of 393 amino acids. The homology and phylogenetic analyses indicate a remarkable similarity between peach PG and other ripening related PG. And seven consensus sequences have revealed in peach PG compared to the PG from other plants. However, the profound divergence with other PG and the unique structure features suggest that peach PG probably belongs to a new evolutionary class. In RT PCR analysis, pMT18 related RNA was undetectable in leaves, and was much abundant in ripe fruits. The ripening specific expression pattern of this cDNA will be useful in investigating the roles of PG in fruit ripening and developing a transgenic peach with the improved post harvesting quality in the future.展开更多
Three endo-polygalacturonases(endoPGs) from a newly isolated Penicillum oxalicum(CGMCC 0907) capable of utilizing waste biomass as growth substrate were separated and purified to homogeneity by ultra-filtration,affini...Three endo-polygalacturonases(endoPGs) from a newly isolated Penicillum oxalicum(CGMCC 0907) capable of utilizing waste biomass as growth substrate were separated and purified to homogeneity by ultra-filtration,affinity adsorption chromatography,CM-cellulose column chromatography,and Sephadex G-100 gel filtration chromatography with the overall yield of 64.5% from the crude enzyme.The specific activities and recovery rates of endoPG-1,endoPG-2 and endoPG-3 were 1120 U/mg and 21.6%,1350 U/mg and 25.9%,and 1560 U/mg and 17.0%,respectively.The three purified endoPGs had a close molecular weight to 41 kDa as estimated by SDS-PAGE.The optimum temperature and pH for the function of them were 65℃ and 5.0,55℃ and 5.0,50℃ and 5.5,respectively.Their pI and Km values were 5.9 and 0.78 mg/mL,6.0 and 1.2 mg/mL,and 6.1 and 2.0 mg/mL,respectively.展开更多
Fruit ripening is a complex process and is regulated by many factors. Ethylene and polygalacturonase （PG）, lipoxygenase （LOX）, expansin （EXP） are all critical regulating factors in fruit ripening and softening p...Fruit ripening is a complex process and is regulated by many factors. Ethylene and polygalacturonase （PG）, lipoxygenase （LOX）, expansin （EXP） are all critical regulating factors in fruit ripening and softening process. With antisense ACS tomato, Nr mutant tomato and cultivated tomato as materials, Northern blot hybridization showed that PG, LeEXP1 and LOXexpressed differently in different parts of cultivated tomato fruit during ripening, which was related to fruit ripening. The ripening process of columella and radial pericarp was faster than pericarp. In both Nr mutant and antisense ACS transgenic tomato fruit, expression levels ofPG, LeEXPI and LOXwere generally lower than those in cultivated fruit but still related to fruit ripening. The expression levels ofPG, LeEXP1 and LOX increased in the mature green tomato fruits after 0.5 h treatment with ethylene （100 μL/L）. These results indicate that gene expression ofPG, LeEXP1 and LOXwere positively regulated by ethylene. The time and cumulative effect of the concentration exists in the expression of PG regulated by ethylene. The regulation of LOX expression mainly depended on the fruit development after great amount of ethylene was produced. PG played a major role in ripening and softening of tomato fruit, and cooperated with the regulation of EXP and LOX.展开更多
基金supported by the Ministerio de Ciencia,Innovación y Universidades and FEDER EU funds(grant numbers AGL2017-86531-C2-1R and PID2020-118468RB-C21)the University of Malaga(grant number B1-2020_09)+1 种基金CS-R was awarded a PhD Fellowship from the Ministerio de Ciencia,Innovación y Universidades(PRE2018-085509)PhD Program Advanced Biotechnology,University of Málaga.
文摘Firmness is one of the most important fruit quality traits in strawberries.The postharvest shelf life of this soft fruit is highly limited by the loss of firmness,where cell wall disassembly plays an important role.Previous studies demonstrated that the polygalacturonase FaPG1 has a key role in remodelling pectins during strawberry softening.In this study,FaPG1 knockout strawberry plants have been generated using the CRISPR/Cas9 system delivered via Agrobacterium tumefaciens.Ten independent lines,cv.“Chandler”,were obtained,and all of them were successfully edited as determined by PCR amplification and T7 endonuclease assay.The targeted mutagenesis insertion and deletion rates were analyzed using targeted deep sequencing.The percentage of edited sequences varied from 47%up to almost 100%,being higher than 95%for seven of the selected lines.Phenotypic analyses showed that 7 out of the eight lines analyzed produced fruits significantly firmer than the control,ranging from 33 to 70%increase in firmness.There was a positive relationship between the degree of FaPG1 editing and the rise in fruit firmness.Minor changes were observed in other fruit quality traits,such as colour,soluble solids,titratable acidity or anthocyanin content.Edited fruits showed a reduced softening rate during postharvest,displayed a reduced transpirational water loss,and were less damaged by Botrytis cinerea inoculation.The analysis of four potential off-target sites revealed no mutation events.In conclusion,editing the FaPG1 gene using the CRISPR/Cas9 system is an efficient method for improving strawberry fruit firmness and shelf life.
基金Supported by Natural Science Foundation of Yunnan Province(2011FB049)National Natural Science Foundation of China(31260481,31460516)+1 种基金Fund of Yunnan Education Department(2013Y251)Development Fund of the Department of Life Science and Technology,Kunming University(GXKM201505)
文摘In this study, 1 500 bp PG gene promoter was amplified from leaves of tomato cultivar ＇ Zhongshu No. 4＇. The bioinformatic analysis of PG promoter sequences was conducted. PG promoter elements were predicted and analyzed by PLANTCARE and PLACE. The results showed that tomato PG promoter contained multiple c/s-acting regulatory elements such as typical basic elements TATA-Box and CAAT-Box, light responsive elements 3-AF1 binding site, ATl-motif, ATCT- motif, Box 4, Box I, GA-motif, GTl-motif, Spl and MRE, heat stress-responsive element HSE, ethylene-responsive element ERE, meristem-specifie regulatory element CCGTCC-box, endosperm expression-related regulatory elements GCN4_motif and Skn-l_motif, defense and stress responsiveness element TC-rich repeats, and circadian control-related element circadian, indicating that the expression of tomato PG gene is related to light, temperature, hormone, stress and other factors. This study laid the foundation for subsequent research about regulation of PG gene expression in plants.
文摘目的:探讨腹腔镜胃癌D2根治术患者经协同护理及患者自评-主观全面评定(patient-generated subjective global assessment,PG-SGA)个体化营养干预的效果。方法:选择蚌埠医科大学第一附属医院2020年8月—2021年11月收治的107例腹腔镜胃癌D2根治术患者,随机分为参照组(n=54,行常规护理)和试验组(n=53,行协同护理+基于PG-SGA个体化营养干预)。对比两组患者的心理状态及生活质量。结果:试验组干预后汉密尔顿焦虑量表(Hamilton anxiety scale,HAMA)、汉密尔顿抑郁量表(Hamilton depression scale,HAMD)评分均较参照组低,健康调查简表(the MOS item short form health survey,SF-36)各维度评分均较参照组高,差异均有统计学意义(P<0.05)。结论:协同护理联合基于PG-SGA个体化营养干预应用于腹腔镜胃癌D2根治术患者,能够有效改善患者的心理状态,提升其生活质量。
文摘Polygalacturonase (PG,EC3.2.1.15) is the key cell wall hydrolase in fruit ripening. The identification and characterization of a full length cDNA (pMT18) encoding for PG from Feicheng peach (Prunus persica (L.) Batsch cv. Feicheng) is described. The pMT18 clone is 1188 bp in length, with an open reading frame of 393 amino acids. The homology and phylogenetic analyses indicate a remarkable similarity between peach PG and other ripening related PG. And seven consensus sequences have revealed in peach PG compared to the PG from other plants. However, the profound divergence with other PG and the unique structure features suggest that peach PG probably belongs to a new evolutionary class. In RT PCR analysis, pMT18 related RNA was undetectable in leaves, and was much abundant in ripe fruits. The ripening specific expression pattern of this cDNA will be useful in investigating the roles of PG in fruit ripening and developing a transgenic peach with the improved post harvesting quality in the future.
基金Supported by National Natural Science Foundation of China (No. 30600082)National Key Technology R&D Program of China (No. 2008BADA7B01)Beijing Municipal Commission of Education (No. KM200811417006)
文摘Three endo-polygalacturonases(endoPGs) from a newly isolated Penicillum oxalicum(CGMCC 0907) capable of utilizing waste biomass as growth substrate were separated and purified to homogeneity by ultra-filtration,affinity adsorption chromatography,CM-cellulose column chromatography,and Sephadex G-100 gel filtration chromatography with the overall yield of 64.5% from the crude enzyme.The specific activities and recovery rates of endoPG-1,endoPG-2 and endoPG-3 were 1120 U/mg and 21.6%,1350 U/mg and 25.9%,and 1560 U/mg and 17.0%,respectively.The three purified endoPGs had a close molecular weight to 41 kDa as estimated by SDS-PAGE.The optimum temperature and pH for the function of them were 65℃ and 5.0,55℃ and 5.0,50℃ and 5.5,respectively.Their pI and Km values were 5.9 and 0.78 mg/mL,6.0 and 1.2 mg/mL,and 6.1 and 2.0 mg/mL,respectively.
基金Supported by National Project of Scientific and Technical Supporting Programs Funded by Ministry of Science and Technology of China (No.2006BAD22B01)National Natural Science Foundation of China (No.30800767)Postdoctoral Fund of China (No.20080430725)
文摘Fruit ripening is a complex process and is regulated by many factors. Ethylene and polygalacturonase （PG）, lipoxygenase （LOX）, expansin （EXP） are all critical regulating factors in fruit ripening and softening process. With antisense ACS tomato, Nr mutant tomato and cultivated tomato as materials, Northern blot hybridization showed that PG, LeEXP1 and LOXexpressed differently in different parts of cultivated tomato fruit during ripening, which was related to fruit ripening. The ripening process of columella and radial pericarp was faster than pericarp. In both Nr mutant and antisense ACS transgenic tomato fruit, expression levels ofPG, LeEXPI and LOXwere generally lower than those in cultivated fruit but still related to fruit ripening. The expression levels ofPG, LeEXP1 and LOX increased in the mature green tomato fruits after 0.5 h treatment with ethylene （100 μL/L）. These results indicate that gene expression ofPG, LeEXP1 and LOXwere positively regulated by ethylene. The time and cumulative effect of the concentration exists in the expression of PG regulated by ethylene. The regulation of LOX expression mainly depended on the fruit development after great amount of ethylene was produced. PG played a major role in ripening and softening of tomato fruit, and cooperated with the regulation of EXP and LOX.
