基因工程技术在甜瓜属作物分子育种中的应用与发展

黄瓜和甜瓜均属于葫芦科甜瓜属作物,作为重要的园艺作物在全世界广泛种植。我国黄瓜和甜瓜的种植面积与产量均居世界第一。不断培育优质高抗的新品种是促进产业持续发展的重要推动力,但常规育种存在转育效率低、育种周期长等问题。多年来瓜类育种家们一直致力于甜瓜属作物基因工程育种方法的探索和研究,随着黄瓜和甜瓜在基因组研究领域取得了重大突破,大量重要功能基因的发掘与克隆为基因工程育种带来了机遇,也促使甜瓜属作物基因工程技术研究进入了快速发展的阶段。就基因工程技术在甜瓜属作物分子遗传育种研究中的应用和发展现状、存在问题以及未来发展趋势等进行了探讨。     

我国甜瓜属蔬菜作物特异基因资源的挖掘和利用

对我国甜瓜属蔬菜作物基因资源的优势和基因资源研究利用中存在的问题进行了阐述,提出了当前我国甜瓜属特异基因资源发掘研究的工作重点。     

外源DNA导入与食用菌育种

外源DNA的导入即提取供体的DNA,把它导入受体,使一定异源DNA片段整合到染色体上的一种基因转化手段。食用菌利用外源DNA导入进行基因转化是一种较先进又有效的育种手段,它可以克服原生质体融合难以得到融合于的困难,又不需要一定载体及限制性内切酶,也能有效地克服种属间远缘杂交的困难。外源DNA导入的方法有: 1.显微注射法:用注射微量吸管注射外源物质到细胞或原生质体中。将注射后的细胞或原生质体进行培养,最后用遗传标志成分子杂交等方法分析外源基因是否导入。 Hammer以微注射外源基因组整合到大型家畜基因中,并得到表达。Marikawa微管注射固定的烟草叶肉原生质体中并整合到基因组中,     

发根农杆菌介导甜瓜转基因过表达体系的建立

遗传转化是实现基因功能验证和基因快速转育的重要手段,但目前甜瓜的遗传转化依旧存在众多技术难题。笔者利用野生型发根农杆菌K599侵染甜瓜'E31',成功诱导甜瓜产生了不定根。借助该技术,将含有GUS基因和EGF基因的过表达载体pCAMBIA3301和pCAMBIA1300-ProSuper导入发根农杆菌K599,利用转化后的农杆菌侵染甜瓜'E31',成功诱导出了不定根。PCR检测结果表明,新生不定根的阳性率达到100%。通过GOS染色和激光共聚焦显微镜检测,在不定根中检测到GOS基因和EGFP基因的大量表达,成功实现了外源基因在甜瓜新生不定根内过表达。该方法周期短、操作方便、转化率高,可实现基因功能的快速验证,为甜瓜根系抗病抗逆方面的研究提供技术支持。     

甘薯种质资源的创新与利用

甘薯为无性繁殖作物，种质资源材料的创新可通过3种途径实现，常规杂交为主要应用方法；通过无性系变异选择获得特定变异的种质材料有一定难度；通过原生质体融合导入近缘野生种的基因，或通过转基因技术导入外源基因，生物技术的应用在甘薯上取得了一些进展。目前甘薯的主要用途分为两大类，一是利用薯肉为桔红心和紫心的品种作为保健型鲜食品种，二是利用高产、高淀粉含量的淀粉型品种作为工业原料。     

薄皮甜瓜白啄瓜枯萎病抗性基因Fom-2的同源克隆与表达分析

枯萎病是甜瓜等瓜类作物的毁灭性病害，Fom-2基因介导了甜瓜抗枯萎病生理小种0和1，抗枯萎病育种是甜瓜等瓜类作物育种的重要目标之一。通过同源克隆方法从薄皮甜瓜白啄瓜中克隆到了枯萎病抗性基因Fom-2，推定编码1099个氨基酸，所推定的蛋白质序列经Blast分析与数据库中NBS-LRR类R基因具有较高的同源性。进一步序列...     

花粉管通道导入外源DNA方法的研究

在总结前人研究的基础上,认为花粉管通道法导入外源DNA的方法可分为花粉粒携带法、子房注射法和柱头切除法。3种导入外源基因的方法已在植物转基因育种研究中被广泛运用。在实际运用中应针对某一特定的受体植物,根据受体植物的雌蕊结构特点选择合适的导入外源基因的方法,可提高花粉管通道法的转化效果和转化率。     

黄瓜与酸黄瓜异源多倍体3种同工酶表达基因组剂量效应

 以甜瓜属（Cucumis）黄瓜与酸黄瓜的3种不同倍性异源多倍体 - 双单倍体种间杂种F₁（2n = 2x = 19,HC）、异源三倍体黄瓜（2n = 3x = 26,HCC）和异源四倍体C. hytivus (2n = 4x = 38,HHCC)及其双亲为试材,比较研究过氧化物酶（POD）、谷氨酸脱氢酶（GDH）和天冬氨酸转氨酶（AAT）在不同倍性异源多倍体中的酶谱表达特性,探讨基因组剂量对酶谱表达的影响。结果表明：不同倍性异源多倍体的酶带染色力表现出明显的差异,但酶带数和酶带分布基本一致。在AAT同工酶中,酶带的表达随基因组剂量的提高而增强,在POD和GDH同工酶中,大部分酶带的表达随基因组剂量的提高而减弱。这些结果表明甜瓜属异源多倍体在不同的酶系统中,酶谱表达可能会表现出截然相反的基因组剂量效应。本研究结果可为     

6 - 磷酸葡萄糖酸脱氢酶基因(6PGDH) 的克隆与甜瓜属异源四倍体的分子验证

 根据已发表的黄瓜6 - 磷酸葡萄糖酸脱氢酶(6-phosphogluconate dehydrogenase ) 基因6PGDH的cDNA序列(GenBank登录号: EU815934) 设计引物, 扩增了甜瓜属异源四倍体新种及野生种和栽培黄瓜‘北京截头’的6PGDH基因片段, 测序结果表明3个种的6PGDH基因片段序列表现出高度的一致性,长度均为1 098 bp包含936 bp编码311个氨基酸的阅读框和162 bp的3′非翻译区末端, 片段内无内含子存在。利用DNAMAN软件检测到异源四倍体新种与野生种和栽培黄瓜‘北京截头’氨基酸的差异位点有3个, 碱基差异位点有22个, 差异位点的遗传分布符合孟德尔遗传学定律, 从而在分子水平证实了异源四倍体新种是野生种和栽培黄瓜‘北京截头’的杂交后代。     

甜瓜属REMAP分子标记体系的建立及应用

基于甜瓜属Ty1-copia类逆转座子的长末端重复序列信息,结合ISSR引物序列,在体系优化的基础上建立了适用于甜瓜属物种的REMAP(retrotransposon-microsatellite amplified polymorphism)标记体系。对46份甜瓜属不同类型材料进行聚类分析,证明该标记体系能有效检测甜瓜属不同类型材料间的多态性,可以用于甜瓜属作物品种鉴定及指纹图谱构建。     

根癌农杆菌介导的白菜类作物转化体系研究进展

白菜类作物是我国乃至世界性的大众蔬菜,利用农杆菌介导技术将目的基因转入蔬菜中,从而选育具有特殊性状的新品种。文章综述了根癌农杆菌介导的白菜类作物高频再生转化体系构建的影响因素,简要介绍了转化植株的分子检测及遗传行为,在此基础上分析了目前遗传转化技术在白菜类作物改良上存在的问题及广阔的应用前景。     

金塔地区外来植物调查研究

通过样方法和样线法对金塔地区外来植物进行了实地调查。结果表明:该地区共有外来植物41科116属152种,其中食用、观赏和绿化种类分别占38%、26%和22%。金塔地区的外来植物集中分布在人口密集的城镇及人类活动频繁的绿洲及其周边地区,外来植物的引入丰富了金塔地区的物种多样性,促进了地方经济发展和生态环境建设,但少数杂草类入侵物种的生态危害也很明显。     

转基因抗病毒病四倍体西瓜的培育

利用西瓜花叶病毒2号(WMV-2)外壳蛋白(CP)基因、小西葫芦黄花叶病毒(ZYMV)复制酶(NIb)基因和黄瓜花叶病毒(CMV)复制酶基因构建三价基因的植物表达载体,通过农杆菌介导转化四倍体西瓜植株,经分子检测证明目的基因成功地导入西瓜植株,并在后代中稳定遗传。经温室及大田接种鉴定,T3代转基因西瓜抗病毒性达中抗水平,为抗病毒无籽西瓜新品种的培育提供了可能性。     

芸薹属作物异附加系的研究进展

 综述了芸薹属作物异附加系的培育概况、鉴定方法及应用前景,分析讨论了存在的问题及今后的发展方向。     

Road verges as invasion corridors? A spatial hierarchical test in an arid ecosystem

Disturbed habitats are often swiftly colonized by alien plant species. Human inhabited areas may act as sources from which such aliens disperse, while road verges have been suggested as corridors facilitating their dispersal. We therefore hypothesized that (i) houses and urban areas are propagule sources from which aliens disperse, and that (ii) road verges act as corridors for their dispersal. We sampled presence and cover of aliens in 20 plots (6 × 25 m) per road at 5-km intervals for four roads, nested within three localities around cities (n = 240). Plots consisted of three adjacent nested transects. Houses (n = 3,349) were mapped within a 5-km radius from plots using topographical maps. Environmental processes as predictors of alien composition differed across spatial levels. At the broadest scale road-surface type, soil type, and competition from indigenous plants were the strongest predictors of alien composition. Within localities disturbance-related variables such as distance from dwellings and urban areas were associated with alien composition, but their effect differed between localities. Within roads, density and proximity of houses was related to higher alien species richness. Plot distance from urban areas, however, was not a significant predictor of alien richness or cover at any of the spatial levels, refuting the corridor hypothesis. Verges hosted but did not facilitate the spread of alien species. The scale dependence and multiplicity of mechanisms explaining alien plant communities found here highlight the importance of considering regional climatic gradients, landscape context and road-verge properties themselves when managing verges. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

大白菜-结球甘蓝单体异附加系试管苗生根能力的研究

以大白菜-结球甘蓝系列单体异附加系试管苗为试材,比较了不同单体异附加系试管苗的生根率和移栽成活率,研究了不同浓度NAA对单体异附加系试管苗生根和根尖细胞分裂的影响。结果表明：AA+C1、AA+C2、AA+C3、AA+C4、AA+C5和AA+C6的试管苗生根较好,移栽成活率高,AA+C7、AA+C8和AA+C9的试管苗不易生根,移栽成活率低;1/2MS培养基附加1.5 mg?mL-1 NAA能有效诱导试管苗生根,提高试管苗生根质量和移栽成活率。     

Landscape fragmentation,land-use legacy and propagule pressure promote plant invasion on coastal dunes: a patch-based approach

Coastal dunes and sand areas are reported to be among the habitats most invaded by alien species in Europe. Landscape pattern could be a significant driver in invasion processes in parallel with land-use legacy. Fragmentation of natural habitats combined with the availability of propagules from the surrounding matrix may enhance the invisibility of ecological communities. Based on multitemporal land cover maps (1954–2008) and a floristic database, we analyzed how habitat fragmentation, propagule pressure and land-use legacy have affected alien plants’ presence and richness on natural dune patches along the Lazio Coast (Central Italy). Floristic data were derived from an existing geo-database of random vegetation plots (64 m²). A set of landscape patch-based metrics, considered to be adequate proxies of the main processes affecting alien invasion and richness, was calculated. First, we fit a generalized linear model (GLM) with binomial errors to assess which landscape metrics are influencing patch invasion. Second, we extracted invaded patches and, with GLMs, we investigated how landscape metrics affect average alien species richness. Alien invasion and alien richness seem to be affected by different processes: although alien invasion of each patch is strongly associated with its land-use legacy, the richness of aliens is more affected by landscape fragmentation and by the propagule pressure to which patch is exposed. By integrating spatial and temporal landscape metrics with floristic data, we were able to disentangle the relations of landscape fragmentation, propagule pressure and land-use legacy with the presence and richness of alien plants. The methodological approach here adopted could be easily extended to other alien species and ecosystems, offering scientifically sound support to prevent the high economic costs derived from both the control and the eradication of aliens.     

附加甘蓝3号染色体的大白菜单体异附加系的获得与研究

 以大白菜—结球甘蓝异源三倍体杂交种（AAC）与二倍体大白菜（AA）的回交一代为材料，利用根尖体细胞核型分析法鉴定出大白菜—结球甘蓝3号单体异附加系；利用甘蓝连锁群特异SSR标记鉴定表明，该单体异附加系中的外源染色体与甘蓝的03连锁群相对应；该单体异附加系植株矮小，花枝细弱倒伏，花粉整齐度、花粉生活力和结籽率均较低，花粉母细胞减数分裂过程中存在染色体不均衡分离及染色体落后等现象。     

Incidence and intensity of tree infestation by the mistletoe Erianthemum dregei (Eckl. & Zeyh.) V. Tieghem in Durban,South Africa

Relatively few studies have investigated mistletoe infestation of tree species in urban settings, and even fewer considered infestation of alien tree species. Here, mistletoe infestation incidence and intensity were studied in Durban, South Africa. The relationship of infestation to tree CBH and height was examined in detail for the mistletoe species Erianthemum dregei. Although E. dregei is known to have a wide range of host taxa, we report E. dregei infestation on 27 new host species for the first time. E. dregei occurred on 30 host species with a preference for the alien host tree Melia azedarach in our study area. The relationship between infestation and CBH indicated that the trees (irrespective of species) of ≥200 cm were more susceptible to the mistletoe attack. The implications of this pattern, as well as those of high host diversity and especially of infestation of alien trees are discussed in a conservation perspective.     

大白菜—结球甘蓝4号染色体二体异附加系的获得与性状分析

以大白菜—结球甘蓝4号染色体单体异附加系自交后代为材料,通过根尖染色体计数法鉴定,从36株中筛选出3株2n = 22的植株。染色体核型分析结果表明,该3株为大白菜—结球甘蓝4号染色体二体异附加系。该二体异附加系在生殖生长时期表现植株较矮小,基生叶卵圆形、绿色,茎生叶披针形、浅绿色,开花时间晚,黄花,花器官较大白菜—结球甘蓝4号染色体单体异附加系小,花粉量少、结荚率和结籽率低。减数分裂观察发现：后期Ⅰ染色体主要以11︰11分离方式为主,所占比率为70.59%;后期Ⅱ染色体以11︰11︰11︰11为主,其比率为69.57%。