西兰花采后黑斑病的鉴定以及致病机理的研究
摘要
西兰花(Brassica oleacea L.var.italica)营养价值丰富而且具有公认的抗癌功效。但是在采后阶段,它会受到各种病原菌的侵染并且迅速衰老黄化,失去原有的商品性。对引起西兰花采后病害的病原进行分离鉴定,进而深入研究其致病机理可为今后制定控制措施提供理论依据。本研究对引起西兰花采后黑斑病的主要病原真菌进行了形态学和分子生物学的鉴定,并对该菌促进西兰花衰老黄化的机理进行了探讨。研究结果表明:
1.对上市销售的西兰花的调查发现,黑斑病是西兰花销售过程中最常见的采后病害之一,西兰花的采后黑斑病往往与其花体的衰老相伴,感染黑斑病的花球会快速衰老黄化,失去商品性。
2.根据柯赫氏法则,对分离自西兰花黑斑病的病原进行致病性鉴定的结果表明,导致西兰花采后病害的主要病原为链格孢属真菌,其中菌株HLYB-4为引起花球黑斑病的主要病原菌,该菌的菌落呈青褐色,分生孢子青褐色、手雷状呈链状排列,大小约为24～51.5×7.5～23μm,2～6个横隔,0～3个纵隔,喙部不发达,为单细胞假喙,形态学初步鉴定为芸苔生链格孢菌(Alternaria brassicicola)应用rDNA-ITS序列比对法和十字花科链格孢属菌的特异标记引物Abra1和Abra2扩增鉴定进一步将引起西兰花采后黑斑病病原定名为A.brassicicola.
3.对西兰花黑斑病病原A.brassicicola的适宜生长温度和pH的研究表明,该菌生长的最适温度为25℃,最适pH5～8。5℃低温不能完全阻止其菌丝生长和孢子形成,而35℃可有效地抑制该菌在体外和植物体上的生长和产孢;A.brassicicola的分生孢子在pH2～12均可萌发,在pH5～9三小时内萌发率接近100%。此外,50ppm的外源乙烯能够促进其孢子萌发。
4.对接种A.brassicicola西兰花的生理代谢指标测定结果表明,接种第72小时的花球开始黄化,叶绿素迅速降解,乙烯释放量和呼吸速率显著升高,接菌组乙烯释放高峰比对照组提早24小时形成。然而,西兰花采后Vc的迅速下降与病原菌侵染没有显著的对应关系。
5.对西兰花衰老调节基因转录水平的分析表明,接种A.brassicicola后西兰花的ACC氧化酶基因BoACO2比对照组提前24小时上调表达;ACC合成酶基因BoACS2也在接种后的72小时内明显上调;叶绿素分解酶基因BoCLH1在前72小时的转录活性也明显高于对照组,生理生化分析与分子证据证明A.brassicicola是诱导西兰花采后的黄化衰老的重要因素之一。
上述研究结果显示,提高西兰花采后贮藏的品质,延长货架期必须严格控制西兰花的采后黑斑病菌A.brassicicola的发生。
Broccoli(Brassica oleacea L.var.italica) is nutritional and regarded as one anti-cancer vegetable.However,it is vulnerable to various pathogens and senescences rapidly after harvest,which remarkably shortens its market life.So it's meaningful to separate and identify these broccoli post-harvest pathogens so as to make effective control protocol.Our research identified the black spot disease pathogen of post-harvest broccoli with the help of morphological and molecular methods and explored its contribution to the senescence of the plants.The results are as follows,
a.According to our market research,Black spot disease seems to be one of the most common diseases on post-harvest broccoli.This pathogen is also related with the aging of the plants.The infected plants senescenced rapidly and lost its market value.
b.According to Koch's Postulates,pathogens on post-harvest broccoli are identificated as Alternaria spp.And strain HLYB-4 was proved to be the cause of the black spot,which produced a green-brown colony and whose spores are 24 to 51.5×7.5 to 23μm grenade-like with two to six transverse septa,zero to three mediastinal septa and fake rostrum.It was identified as Alternaria brassicicola by morphologically analysis and further proved by the rDNA-ITS blast and particular primers Abral andAbra2.
c.The optimal temperature and pH of A.brassicicola are 25℃and pH 5-8 respectfully.Low storage temperature(5℃) can't block its hypha growth and sporation totally.However,high temperature(35℃) seems to effectively restrain its growth and sporation both in vitro and in vivo.In addition,its spores could germinate from pH 2-12 and the germination rate reached 100%within 3 hours at pH 5-9.Moreover,50ppm exogenous ethylene seems to contribute to its germination.
d.After pathogen inoculating on the healthy head of broccoli,obvious yellowing could be seen within 72 hours with the degradation of its chlorophyll.High ethylene output and respiration rate could be measured while the inoculated group could be tested 24 hours earlier than the control.However,the rapid descendant of Vitamin C seems not to be related with the infection.
e.The RT-PCR result indicates that the transcription of Bo-ACS2 enhanced in the first 72h in the inoculated group and Bo-ACO2 have up-regulated in the first 24 hours in the inoculated group as well,which is one day earlier than the control group.And the transcription of Bo-CLH1 of infected group in the first 72 hours is obviously higher than the control group.So both physiological and molecular results indicated that A.brassicicola induced the senescence of post-harvest broccoli.
Above all,in order to improve the preservation of broccoli and extend its market life,it is critical to prevent the occurrence of black spot caused by A.brassicicola.
1.对上市销售的西兰花的调查发现,黑斑病是西兰花销售过程中最常见的采后病害之一,西兰花的采后黑斑病往往与其花体的衰老相伴,感染黑斑病的花球会快速衰老黄化,失去商品性。
2.根据柯赫氏法则,对分离自西兰花黑斑病的病原进行致病性鉴定的结果表明,导致西兰花采后病害的主要病原为链格孢属真菌,其中菌株HLYB-4为引起花球黑斑病的主要病原菌,该菌的菌落呈青褐色,分生孢子青褐色、手雷状呈链状排列,大小约为24～51.5×7.5～23μm,2～6个横隔,0～3个纵隔,喙部不发达,为单细胞假喙,形态学初步鉴定为芸苔生链格孢菌(Alternaria brassicicola)应用rDNA-ITS序列比对法和十字花科链格孢属菌的特异标记引物Abra1和Abra2扩增鉴定进一步将引起西兰花采后黑斑病病原定名为A.brassicicola.
3.对西兰花黑斑病病原A.brassicicola的适宜生长温度和pH的研究表明,该菌生长的最适温度为25℃,最适pH5～8。5℃低温不能完全阻止其菌丝生长和孢子形成,而35℃可有效地抑制该菌在体外和植物体上的生长和产孢;A.brassicicola的分生孢子在pH2～12均可萌发,在pH5～9三小时内萌发率接近100%。此外,50ppm的外源乙烯能够促进其孢子萌发。
4.对接种A.brassicicola西兰花的生理代谢指标测定结果表明,接种第72小时的花球开始黄化,叶绿素迅速降解,乙烯释放量和呼吸速率显著升高,接菌组乙烯释放高峰比对照组提早24小时形成。然而,西兰花采后Vc的迅速下降与病原菌侵染没有显著的对应关系。
5.对西兰花衰老调节基因转录水平的分析表明,接种A.brassicicola后西兰花的ACC氧化酶基因BoACO2比对照组提前24小时上调表达;ACC合成酶基因BoACS2也在接种后的72小时内明显上调;叶绿素分解酶基因BoCLH1在前72小时的转录活性也明显高于对照组,生理生化分析与分子证据证明A.brassicicola是诱导西兰花采后的黄化衰老的重要因素之一。
上述研究结果显示,提高西兰花采后贮藏的品质,延长货架期必须严格控制西兰花的采后黑斑病菌A.brassicicola的发生。
Broccoli(Brassica oleacea L.var.italica) is nutritional and regarded as one anti-cancer vegetable.However,it is vulnerable to various pathogens and senescences rapidly after harvest,which remarkably shortens its market life.So it's meaningful to separate and identify these broccoli post-harvest pathogens so as to make effective control protocol.Our research identified the black spot disease pathogen of post-harvest broccoli with the help of morphological and molecular methods and explored its contribution to the senescence of the plants.The results are as follows,
a.According to our market research,Black spot disease seems to be one of the most common diseases on post-harvest broccoli.This pathogen is also related with the aging of the plants.The infected plants senescenced rapidly and lost its market value.
b.According to Koch's Postulates,pathogens on post-harvest broccoli are identificated as Alternaria spp.And strain HLYB-4 was proved to be the cause of the black spot,which produced a green-brown colony and whose spores are 24 to 51.5×7.5 to 23μm grenade-like with two to six transverse septa,zero to three mediastinal septa and fake rostrum.It was identified as Alternaria brassicicola by morphologically analysis and further proved by the rDNA-ITS blast and particular primers Abral andAbra2.
c.The optimal temperature and pH of A.brassicicola are 25℃and pH 5-8 respectfully.Low storage temperature(5℃) can't block its hypha growth and sporation totally.However,high temperature(35℃) seems to effectively restrain its growth and sporation both in vitro and in vivo.In addition,its spores could germinate from pH 2-12 and the germination rate reached 100%within 3 hours at pH 5-9.Moreover,50ppm exogenous ethylene seems to contribute to its germination.
d.After pathogen inoculating on the healthy head of broccoli,obvious yellowing could be seen within 72 hours with the degradation of its chlorophyll.High ethylene output and respiration rate could be measured while the inoculated group could be tested 24 hours earlier than the control.However,the rapid descendant of Vitamin C seems not to be related with the infection.
e.The RT-PCR result indicates that the transcription of Bo-ACS2 enhanced in the first 72h in the inoculated group and Bo-ACO2 have up-regulated in the first 24 hours in the inoculated group as well,which is one day earlier than the control group.And the transcription of Bo-CLH1 of infected group in the first 72 hours is obviously higher than the control group.So both physiological and molecular results indicated that A.brassicicola induced the senescence of post-harvest broccoli.
Above all,in order to improve the preservation of broccoli and extend its market life,it is critical to prevent the occurrence of black spot caused by A.brassicicola.
引文
Able A J,Wong L S,Prasad A,O'hare T J.1-MCP is more effective on a floral brassica(Brassica oleracea var.italica L.) than a leafy brassica(Brassica rapa var.chinensis)[J].Postharvest Biology and Technology,2002,26(2):147-155
Ambrosone C B,McCann S E,Freudenheim J L,Marshall J R,Zhang Y S,Shields P G.Breast Cancer Risk in Premenopausal Women Is Inversely Associated with Consumption of Broccoli,a Source of Isothiocyanates,but Is Not Modified by GST Genotype[J].Journal of nutrition,2004,134(5):1134-1138
Andrew M,Peever T L,Pryor B M.An expanded multilocus phylogeny does not resolve morphological species within the small-spored Alternaria species complex[J].Mycologia,2009,101(1):95-109
Azarenko O,Okouneva T,Singletary K W,Jordan M A,Wilson L.Suppression of microtubule dynamic instability and turnover in MCF7 breast cancer cells by sulforaphane[J].Carcinogenesis,2008,29(12):2360-2368
Bartoli C G,Simontacchi M,Guiamet J J,Montaldi E,Puntarulo S.Antioxidant enzymes and lipid peroxidation during aging of Chrysanthemum morifolium RAM petals[J].Plant Science,1995,104(2):161-168
Bell G I.Evolution of simple sequence repeats[J].Computer & Chemistry,1996,20(1):41-48
Bock C H,Thrall P H,Brubaker C L,Burdon J J.Detection of genetic variation in Alternaria brassicicola using AFLP fingerprinting[J].Mycologia,2002,106(4):428-434
Borochov A,Woodson W R.Ph ysiology and biochemistry of flower petal senescence[J].Horticultural Review,1989(11):15-43
Chagu(?) V,Danit L V,Siewers V,Gronover G S,Tudzynski P,Tudzynski B,Sharon A.Ethylene Sensing and Gene Activation in Botrytis cinerea:A Missing Link in Ethylene Regulation of Fungus-Plant Interactions?[J].The American Phytopathological Society,2006,19(1):33-42
常缨,王义权,强胜.链格孢菌菌株致病性及其遗传差异性[J].应用与环境生物 学报,2005,11(4):486-489
陈本懋.维生素C和K_3的抗癌作用[J].生命的化学,1991,11(4):27
陈海平,冯春梅,莫云彬,董荷玲,董国堃.西兰花球茎褐变危害及病原鉴定初报[J].浙江农业科学,2005(1):63-65
陈伟群,张天宇.十字花科上的链格孢属真菌同工酶凝胶电泳分析[J].真菌学报,1994,13(4):295-302
Chiao J W,Chung F L,Kancherla R,Ahmed T,Mittelman A,Conaway C C.Sulforaphane and its metabolite mediate growth arrest and apoptosis in human prostate cancer cells[J].International Journal of Oncology,2002,20(3):631-636
Chou H H,Wu W S.Phylogenetic analysis of internal transcribed spacer regions of the genus Alternaria,and the significance of filament-beaked conidia[J].Mycological Research,2002,106(2):164-169
Cooke D E L,Forster J W,Jenkins P D,Jones D G.Lewis D M.Analysis of intraspecific and interspecific variation in the genus Alternaria by the use of RAPD-PCR[J].Annals of Applied Biology,1998,132(2):197-209
邓俭英,刘忠,康德贤,张玲玲,王绪,方锋学.RFLP 分子标记及其在蔬菜研究中的应用[J].分子植物育种,2005,3(2):245-248
崔香仙.西兰花主要病害及其综合防治[J].特种经济动植物,2005(3):42
Eason J R,Ryan D J,Watson L M,Hedderley D,Christey M C,Braun R H,Coupe S A.Suppression of the cysteine protease,aleutain,delays floret senescence in Brassica oleracea[J].Plant Molecular Biology,2005(57):645-657
Enya J,Togawa M,Takeuchi T,Yoshida S,Tsushima S,Arie T,Sakai T.Biological and phylogenetic characterization of Fusarium oxysporum complex,which causes yellows on Brassica spp.,and proposal of F.oxysporum f.sp rapae,a novel forma specialis pathogenic on B-rapa in Japan[J].Phytopathology,2008,98(4):475-483
韩兆轩,王维民.类黄酮在心血管疾病中的研究进展[J].农产品加工,2007(12):31-33.56
Hardan K,Al-Masri M I,Barakat R.Ethylene production by Botrytis cinerea(causal organism of gray mold disease) and influence of the exogenously applied growth regulators and thier inhibitor on disease development[J].Hebron University Rearch Journal,2005,2(1):55-75
Haristoy X,Angioi-Duprez K,Duprez A,Lozniewski A.Efficacy of Sulforaphane in Eradicating Helicobacter pylori in Human Gastric Xenografts Implanted in Nude Mice[J].American Society for Microbiology,2003,47(12):3982-3984
Hong S G,Maccaroni M,Figuli P J,Pryor B M,Belisario A.Polyphasic classification of Alternaria isolated from hazelnut and walnut fruit in Europe[J].Mycological Research,2006(110):1290-1300
Hong S G,Liu D,Pryor B M.Restriction mapping of the IGS region in Alternaria spp.reveals variable and conserved domains[J].Mycological Research,2005,109(1):87-95
胡廷章,胡宗利,屈霄霄,任彦荣,陈国平.植物脂肪氧化酶的研究进展[J].生物工程学报,2009,25(1):1-9
Irtwange S V.Hot water treatment:A non-chemical alternative in keeping quality during post-harvest handling of citrus fruits[J].Agricultural Engineering International.2006,5(8):1-10
季宇彬,池文杰,邹翔,王志举.西兰花中萝卜硫素提取、分离与抗癌活性研究[J].哈尔滨商业大学学报,2005,21(3):270-273
Kusaba M,Tsuge T.Phylogeny of Alternaria fungi known to produce host-specific toxins on the basis of variation in internal transcribed spacers of ribosomal DNA[J].Current Genetics,1995(28):491-498
Kusaba M,Tsuge T.Coinfection by different isolates of Alternaria alternata in single blank spot lesions of Japanese pear leaves[J].Phytopathology,1994,84(5):447-451
Lemoine M L,Civello P M,Martinez G A,Chaves A R.Effect of combined treatment with hot air and UV-C on senescence and quality parameters of minimally processed broccoli(Brassica oleracea L.var.Italica)[J].Postharvest Biology and Technology,2008,48(1):15-21
李多川,张天宇,吴竞爽.链格孢属(丝孢纲)数值分类研究初探[J].真菌学报,1993,12(3):232-239
李群伟.真菌毒素与人畜健康的研究现状及展望[J].中国预防医学杂志,2004,5(5):409-412 林晓民,李振岐,王少先.真菌rDNA 的特点及在外生菌根菌鉴定中的应用[J].西北农业学报,2005,14(2):120-125
Lina I,Roy W C.Production of Ethylene by Fungi[J].Science,1968,159(3821):1357-1358.
刘少华,陆金萍,朱瑞良,戴富明.一种快速简便的植物病原真菌基因组DNA提取方法[J].植物病理学报,2005,35(4):362-365
刘威,张云川,赵美华,李永军.不同漂烫温度对西兰花品质指标的影响[J].农产品加工,2007(1):86-88
Lund B M,Wyatt G M.Post-harvest Spoilage of Cauliflower by Downy Mildew,Peronospora parasitica[J].Plant Pathology,2007,27(3):143-144
Mercier J,Makhlouf J,Martin R A.Fusarium avenaceum,a pathogen of stored broccoli[J].Canadian Plant Disease Survey,1991,71(2):161-162
Powolny A,Takahashi K,Hopkins R G,Loo G.Induction of GADD gene expression by phenethylisothiocyanate in human colon adenocarcinoma cells[J].Journal of Cellular Biochemistry,2003,90(6):1128-1139
Rao V G.An account of the market and storage diseases of fruits and vegetables in Bombay-Maharashtra[J].Mycopathologia.1966,28(1-2):165-176
Rogiers S Y,Kumar G N M,Knowles N R.Maturation and ripening of fruit of Amelanchier alnifolia Nutt.are accompanied by increasing oxidative stress[J].Annals of Botany,1998(81):203-211
茹水江,王汉荣,王连平,李伟龙,苏英京,黄新灿,金昌林,赵建阳.西兰花菌核病空间分布格局及抽样技术研究[J].浙江农业学报,2006,18(2):63-66
Schouten R E,Zhang X B,Verschoor J A,Otma E C,Tijskens L M M,van Kooten O.Development of colors of broccoli heads as affected by controlled atmosphere storage and temperature[J].Postharvest Biology and Technology,2009(51):27-35
Sharma T R,Tewari J P.RAPD analysis of three Alternaria species pathogenic to Crucifers.Mycological Research,1998,102(7):807-814
沈瑞清,张天宇.培养基对链格孢属(Alternaria Nees)真菌种级形态分类特征影响的研究[J].宁夏农学院学报,2003,24(3):1-10
生吉萍,申琳,罗云波.果蔬成熟和衰老中的重要酶-脂氧合酶[J].果树科学, 1999,16(1):72-77
史国安,郭香凤,张益民,王淑芳.GA3和乙烯利对杏果实采后活性氧代谢的影响[J].园艺学报,1997,24(1):87-88
水果中维生素C的测定[E].http://www.foodmate.net/jianyan/lihua/xiangmu/12097.html
水果、蔬菜制品可滴定酸的测定[E].http://www.peach.net.cn/news/news_show.asp?id=167
Suzuki Y,Uji T,Terai H.Inhibition of senescence in broccoli florets with ethanol vapor from alcohol powder[J].Postharvest Biology and echnology,2004,31(2):177-182
唐晓伟.西兰花营养最高[J].花卉,2006,7:33.
Tian M S,Downs C G,Lill R E,King G A.A role for ethylene in the yellowing of broccoli after harvest[J].Journal of the American Society for Horticultural Science,1994,119(2):276-281
Tohyama A,Tsuda M.Alternaria on cruciferous plants.4.Alternaria species on seed of some cruciferous crops and their pathogenicity[J].Mycoscience,1995(36):257-261
Tran-Dinh Nai,Hocking A.Isolation and characterization of polymorphic microsatellite markers for Alternaria alternate[J].Molecular Ecology Notes,2006(6):405-407
汪恩国,陈克松,李达林.青花菜黑腐病空间分布格局测定及其应用[J].中国植保导刊,2004,24(2):30-31
王贵仁.青花菜的保鲜贮藏[J].蔬菜,2007(8):30
王洪凯,张天宇,张猛.链格孢属真菌分类研究进展[J].山东农业大学学报,2001,32(3):406-410
王洪凯,张天宇,张猛.5.8SrDNA及ITS区序列分析链格孢种级分类[J].菌物系统,2001,20(2):168-173
王洪凯,张天宇.链格孢属小孢子种的RAPD分析[J].菌物系统,2003,22(1):35-41
王晓梅,崔坤,陆艳玲.中国西兰花应用价值及生产、出口前景分析[J].农业信息科学,2008,24(11):478-480.
王志平,刘玉梅,方智远,孙培田,杨丽梅,王小武,庄木,王小琴.青花菜贮藏过程中糖、Vc和蛋白质的变化[J].中国蔬菜,2001(2):27-29
Weir T L,Huff D R,Christ B J,Romaine C R RAPD-PCR analysis of genetic variation among isolates of Alternaria solani and Alternaria alternata from potato and tomato[J].Mycologia,1998,90(5):813-821
肖长坤,李健强,师迎春,郑建秋,Bolkan H.十字花科蔬菜黑斑病菌的PCR鉴定[J],植物病理学报,2005,35(3):278-282
肖长坤,吴学宏,李健强,郑建秋,Hasan B,张文华.白菜黑斑病菌三个种菌株基本培养条件比较[J].菌物学报,2004,23(4):573-579
Yang C Y,Chu F H,Wang Y T,Chen Y T,Yang S F,Shaw J F.Novel broccoli l-aminocyclopropane-l-carboxylate oxidase gene(Bo-ACO3) associated with the late stage of postharvest floret senescence[J].Journal of agricultural and food chemistry,2003,51(9):2569-2575
杨连君.西兰花小档案[J].饮食科学,2003(10):26
叶陈亮,陈伟.青花菜花球衰老进程中呼吸及某些物质代谢的变化[J].园艺学报,1995,22(4):367-371
宇田川俊一,食品のカビ污染と危害[M].日本:幸書房,2004
张敬泽,张天宇,王瑾.链格孢属种间培养性状的分类研究[J].浙江农业大学学报,1997,23(5):511-514
张瑞明.青花菜主要病害发生与防治措施[J].上海蔬菜,2006(2):46-49
张石革.出血与维生素K[J].中国药房,2005,16(24):1920
张天宇.中国真菌志(链格孢属)[M].科学出版社,1998,98-100
张志良,瞿伟菁.植物生理学实验指导[M].北京高等教育出版社(第三版),2003:127-128
朱颖莹,许玲,张晟瑜,刘毅,冯琳燕,李菊芬.进口鲜葡萄市场病害的检测与分析[J].食品科学,2007,28(7):393-396
Ambrosone C B,McCann S E,Freudenheim J L,Marshall J R,Zhang Y S,Shields P G.Breast Cancer Risk in Premenopausal Women Is Inversely Associated with Consumption of Broccoli,a Source of Isothiocyanates,but Is Not Modified by GST Genotype[J].Journal of nutrition,2004,134(5):1134-1138
Andrew M,Peever T L,Pryor B M.An expanded multilocus phylogeny does not resolve morphological species within the small-spored Alternaria species complex[J].Mycologia,2009,101(1):95-109
Azarenko O,Okouneva T,Singletary K W,Jordan M A,Wilson L.Suppression of microtubule dynamic instability and turnover in MCF7 breast cancer cells by sulforaphane[J].Carcinogenesis,2008,29(12):2360-2368
Bartoli C G,Simontacchi M,Guiamet J J,Montaldi E,Puntarulo S.Antioxidant enzymes and lipid peroxidation during aging of Chrysanthemum morifolium RAM petals[J].Plant Science,1995,104(2):161-168
Bell G I.Evolution of simple sequence repeats[J].Computer & Chemistry,1996,20(1):41-48
Bock C H,Thrall P H,Brubaker C L,Burdon J J.Detection of genetic variation in Alternaria brassicicola using AFLP fingerprinting[J].Mycologia,2002,106(4):428-434
Borochov A,Woodson W R.Ph ysiology and biochemistry of flower petal senescence[J].Horticultural Review,1989(11):15-43
Chagu(?) V,Danit L V,Siewers V,Gronover G S,Tudzynski P,Tudzynski B,Sharon A.Ethylene Sensing and Gene Activation in Botrytis cinerea:A Missing Link in Ethylene Regulation of Fungus-Plant Interactions?[J].The American Phytopathological Society,2006,19(1):33-42
常缨,王义权,强胜.链格孢菌菌株致病性及其遗传差异性[J].应用与环境生物 学报,2005,11(4):486-489
陈本懋.维生素C和K_3的抗癌作用[J].生命的化学,1991,11(4):27
陈海平,冯春梅,莫云彬,董荷玲,董国堃.西兰花球茎褐变危害及病原鉴定初报[J].浙江农业科学,2005(1):63-65
陈伟群,张天宇.十字花科上的链格孢属真菌同工酶凝胶电泳分析[J].真菌学报,1994,13(4):295-302
Chiao J W,Chung F L,Kancherla R,Ahmed T,Mittelman A,Conaway C C.Sulforaphane and its metabolite mediate growth arrest and apoptosis in human prostate cancer cells[J].International Journal of Oncology,2002,20(3):631-636
Chou H H,Wu W S.Phylogenetic analysis of internal transcribed spacer regions of the genus Alternaria,and the significance of filament-beaked conidia[J].Mycological Research,2002,106(2):164-169
Cooke D E L,Forster J W,Jenkins P D,Jones D G.Lewis D M.Analysis of intraspecific and interspecific variation in the genus Alternaria by the use of RAPD-PCR[J].Annals of Applied Biology,1998,132(2):197-209
邓俭英,刘忠,康德贤,张玲玲,王绪,方锋学.RFLP 分子标记及其在蔬菜研究中的应用[J].分子植物育种,2005,3(2):245-248
崔香仙.西兰花主要病害及其综合防治[J].特种经济动植物,2005(3):42
Eason J R,Ryan D J,Watson L M,Hedderley D,Christey M C,Braun R H,Coupe S A.Suppression of the cysteine protease,aleutain,delays floret senescence in Brassica oleracea[J].Plant Molecular Biology,2005(57):645-657
Enya J,Togawa M,Takeuchi T,Yoshida S,Tsushima S,Arie T,Sakai T.Biological and phylogenetic characterization of Fusarium oxysporum complex,which causes yellows on Brassica spp.,and proposal of F.oxysporum f.sp rapae,a novel forma specialis pathogenic on B-rapa in Japan[J].Phytopathology,2008,98(4):475-483
韩兆轩,王维民.类黄酮在心血管疾病中的研究进展[J].农产品加工,2007(12):31-33.56
Hardan K,Al-Masri M I,Barakat R.Ethylene production by Botrytis cinerea(causal organism of gray mold disease) and influence of the exogenously applied growth regulators and thier inhibitor on disease development[J].Hebron University Rearch Journal,2005,2(1):55-75
Haristoy X,Angioi-Duprez K,Duprez A,Lozniewski A.Efficacy of Sulforaphane in Eradicating Helicobacter pylori in Human Gastric Xenografts Implanted in Nude Mice[J].American Society for Microbiology,2003,47(12):3982-3984
Hong S G,Maccaroni M,Figuli P J,Pryor B M,Belisario A.Polyphasic classification of Alternaria isolated from hazelnut and walnut fruit in Europe[J].Mycological Research,2006(110):1290-1300
Hong S G,Liu D,Pryor B M.Restriction mapping of the IGS region in Alternaria spp.reveals variable and conserved domains[J].Mycological Research,2005,109(1):87-95
胡廷章,胡宗利,屈霄霄,任彦荣,陈国平.植物脂肪氧化酶的研究进展[J].生物工程学报,2009,25(1):1-9
Irtwange S V.Hot water treatment:A non-chemical alternative in keeping quality during post-harvest handling of citrus fruits[J].Agricultural Engineering International.2006,5(8):1-10
季宇彬,池文杰,邹翔,王志举.西兰花中萝卜硫素提取、分离与抗癌活性研究[J].哈尔滨商业大学学报,2005,21(3):270-273
Kusaba M,Tsuge T.Phylogeny of Alternaria fungi known to produce host-specific toxins on the basis of variation in internal transcribed spacers of ribosomal DNA[J].Current Genetics,1995(28):491-498
Kusaba M,Tsuge T.Coinfection by different isolates of Alternaria alternata in single blank spot lesions of Japanese pear leaves[J].Phytopathology,1994,84(5):447-451
Lemoine M L,Civello P M,Martinez G A,Chaves A R.Effect of combined treatment with hot air and UV-C on senescence and quality parameters of minimally processed broccoli(Brassica oleracea L.var.Italica)[J].Postharvest Biology and Technology,2008,48(1):15-21
李多川,张天宇,吴竞爽.链格孢属(丝孢纲)数值分类研究初探[J].真菌学报,1993,12(3):232-239
李群伟.真菌毒素与人畜健康的研究现状及展望[J].中国预防医学杂志,2004,5(5):409-412 林晓民,李振岐,王少先.真菌rDNA 的特点及在外生菌根菌鉴定中的应用[J].西北农业学报,2005,14(2):120-125
Lina I,Roy W C.Production of Ethylene by Fungi[J].Science,1968,159(3821):1357-1358.
刘少华,陆金萍,朱瑞良,戴富明.一种快速简便的植物病原真菌基因组DNA提取方法[J].植物病理学报,2005,35(4):362-365
刘威,张云川,赵美华,李永军.不同漂烫温度对西兰花品质指标的影响[J].农产品加工,2007(1):86-88
Lund B M,Wyatt G M.Post-harvest Spoilage of Cauliflower by Downy Mildew,Peronospora parasitica[J].Plant Pathology,2007,27(3):143-144
Mercier J,Makhlouf J,Martin R A.Fusarium avenaceum,a pathogen of stored broccoli[J].Canadian Plant Disease Survey,1991,71(2):161-162
Powolny A,Takahashi K,Hopkins R G,Loo G.Induction of GADD gene expression by phenethylisothiocyanate in human colon adenocarcinoma cells[J].Journal of Cellular Biochemistry,2003,90(6):1128-1139
Rao V G.An account of the market and storage diseases of fruits and vegetables in Bombay-Maharashtra[J].Mycopathologia.1966,28(1-2):165-176
Rogiers S Y,Kumar G N M,Knowles N R.Maturation and ripening of fruit of Amelanchier alnifolia Nutt.are accompanied by increasing oxidative stress[J].Annals of Botany,1998(81):203-211
茹水江,王汉荣,王连平,李伟龙,苏英京,黄新灿,金昌林,赵建阳.西兰花菌核病空间分布格局及抽样技术研究[J].浙江农业学报,2006,18(2):63-66
Schouten R E,Zhang X B,Verschoor J A,Otma E C,Tijskens L M M,van Kooten O.Development of colors of broccoli heads as affected by controlled atmosphere storage and temperature[J].Postharvest Biology and Technology,2009(51):27-35
Sharma T R,Tewari J P.RAPD analysis of three Alternaria species pathogenic to Crucifers.Mycological Research,1998,102(7):807-814
沈瑞清,张天宇.培养基对链格孢属(Alternaria Nees)真菌种级形态分类特征影响的研究[J].宁夏农学院学报,2003,24(3):1-10
生吉萍,申琳,罗云波.果蔬成熟和衰老中的重要酶-脂氧合酶[J].果树科学, 1999,16(1):72-77
史国安,郭香凤,张益民,王淑芳.GA3和乙烯利对杏果实采后活性氧代谢的影响[J].园艺学报,1997,24(1):87-88
水果中维生素C的测定[E].http://www.foodmate.net/jianyan/lihua/xiangmu/12097.html
水果、蔬菜制品可滴定酸的测定[E].http://www.peach.net.cn/news/news_show.asp?id=167
Suzuki Y,Uji T,Terai H.Inhibition of senescence in broccoli florets with ethanol vapor from alcohol powder[J].Postharvest Biology and echnology,2004,31(2):177-182
唐晓伟.西兰花营养最高[J].花卉,2006,7:33.
Tian M S,Downs C G,Lill R E,King G A.A role for ethylene in the yellowing of broccoli after harvest[J].Journal of the American Society for Horticultural Science,1994,119(2):276-281
Tohyama A,Tsuda M.Alternaria on cruciferous plants.4.Alternaria species on seed of some cruciferous crops and their pathogenicity[J].Mycoscience,1995(36):257-261
Tran-Dinh Nai,Hocking A.Isolation and characterization of polymorphic microsatellite markers for Alternaria alternate[J].Molecular Ecology Notes,2006(6):405-407
汪恩国,陈克松,李达林.青花菜黑腐病空间分布格局测定及其应用[J].中国植保导刊,2004,24(2):30-31
王贵仁.青花菜的保鲜贮藏[J].蔬菜,2007(8):30
王洪凯,张天宇,张猛.链格孢属真菌分类研究进展[J].山东农业大学学报,2001,32(3):406-410
王洪凯,张天宇,张猛.5.8SrDNA及ITS区序列分析链格孢种级分类[J].菌物系统,2001,20(2):168-173
王洪凯,张天宇.链格孢属小孢子种的RAPD分析[J].菌物系统,2003,22(1):35-41
王晓梅,崔坤,陆艳玲.中国西兰花应用价值及生产、出口前景分析[J].农业信息科学,2008,24(11):478-480.
王志平,刘玉梅,方智远,孙培田,杨丽梅,王小武,庄木,王小琴.青花菜贮藏过程中糖、Vc和蛋白质的变化[J].中国蔬菜,2001(2):27-29
Weir T L,Huff D R,Christ B J,Romaine C R RAPD-PCR analysis of genetic variation among isolates of Alternaria solani and Alternaria alternata from potato and tomato[J].Mycologia,1998,90(5):813-821
肖长坤,李健强,师迎春,郑建秋,Bolkan H.十字花科蔬菜黑斑病菌的PCR鉴定[J],植物病理学报,2005,35(3):278-282
肖长坤,吴学宏,李健强,郑建秋,Hasan B,张文华.白菜黑斑病菌三个种菌株基本培养条件比较[J].菌物学报,2004,23(4):573-579
Yang C Y,Chu F H,Wang Y T,Chen Y T,Yang S F,Shaw J F.Novel broccoli l-aminocyclopropane-l-carboxylate oxidase gene(Bo-ACO3) associated with the late stage of postharvest floret senescence[J].Journal of agricultural and food chemistry,2003,51(9):2569-2575
杨连君.西兰花小档案[J].饮食科学,2003(10):26
叶陈亮,陈伟.青花菜花球衰老进程中呼吸及某些物质代谢的变化[J].园艺学报,1995,22(4):367-371
宇田川俊一,食品のカビ污染と危害[M].日本:幸書房,2004
张敬泽,张天宇,王瑾.链格孢属种间培养性状的分类研究[J].浙江农业大学学报,1997,23(5):511-514
张瑞明.青花菜主要病害发生与防治措施[J].上海蔬菜,2006(2):46-49
张石革.出血与维生素K[J].中国药房,2005,16(24):1920
张天宇.中国真菌志(链格孢属)[M].科学出版社,1998,98-100
张志良,瞿伟菁.植物生理学实验指导[M].北京高等教育出版社(第三版),2003:127-128
朱颖莹,许玲,张晟瑜,刘毅,冯琳燕,李菊芬.进口鲜葡萄市场病害的检测与分析[J].食品科学,2007,28(7):393-396