Complete chloroplast genomes of Saccharum spontaneum,Saccharum officinarum and Miscanthus floridulus (Panicoideae: Andropogoneae) reveal the plastid view on sugarcane origins

Sugarcane (Saccharum hybrid cultivar) ranks among the world's top 10 food crops and annually provides 60–70% of the sugar produced worldwide. Despite its economic importance there has been no large-scale systematics study of genus Saccharum and the existing model of sugarcane origins has remained largely unchallenged for almost 50 years. For the first time, we have assembled the complete plastid genomes of Miscanthus floridulus (first report for this genus), Saccharum spontaneum and Saccharum officinarum allowing us to elucidate the phylogenetic origins of Saccharum s.s. species. We demonstrate that Saccharum s.s. is divided into four species, with S. spontaneum diverging from the remainder of the genus about 1.5 million years ago and S. robustum diverging 750,000 years ago. Two separate lineages, one leading to S. officinarum and the other leading to modern hybrid cultivars diverged from S. robustum 640,000 years ago. These findings overturn all previous hypotheses on sugarcane origins, demonstrating that sugarcane's antecedents could not have arisen by human action. All modern cultivars share a common Polynesian origin, whereas Old World canes, S. barberi and S. sinense, cluster as a distinct S. officinarum lineage. This makes modern cultivars a distinct species of genus Saccharum, and we formally propose the name Saccharum cultum for the ancestor of all lineages currently classified as Saccharum hybrid cultivars.     

甘蔗细茎野生种云南不同生态类型的RAPD分析

利用25个随机引物对来自云南不同生态类型的82份甘蔗细茎野生种（Saccharnm spontanenm L．）和4份国外种材料进行RAPD标记,结果表明：云南甘蔗细茎野生种不同生态类型的遗传变异较大,具有丰富的遗传多样性,低续度类型的遗传多样性明显高于高纬度类型,在相同的纬度范围内,随着海拔的升高,其多态性逐渐减少,基于分子聚类分析,86份材料被划分为8个不同群体,表现出明显的地理分布的特点,结果初步证明了云南甘蔗细茎野生种可能起源于云南南部低海拔,低纬度地区,而后逐渐向高海拔,高纬度的西北和东北部演化,扩散,提出了云南南部可能是野生甘蔗起源中心之一的观点。     

甘蔗不同部位的固氮酶活性检测

比较巴西2个固氮甘蔗品种‘B1’、‘B8’的叶片、主脉、叶鞘、茎、根和根际周围固氮酶活性的结果表明:茎、根、根际周围的固氮酶活性较高。     

11份割手密遗传多样性的SSR分析

利用10对多态性丰富的SSR引物,以国家甘蔗种质资源圃中保存的14份具有代表性的割手密为对照,对未收集过的云南地区11份割手密（Saccharum spontaneum L.）野生资源进行多样性分析。结果共扩增出233条DNA谱带,与对照相比,新采集材料的多态性条带为207条,其中14条为特有条带,多态性条带比率为0.89。遗传相似性系数和UPGMA聚类分析表明,新采集的材料并没有单独聚为一类,而是比较分散,在相似性系数为0.64处做切割线,参试材料可分为三个类群：第一类群主要由龙门割手密、河边村割手密和福建仙游1号组成;第二类群中包含19份材料,其中新采集的样品有上岗割手密、他拉割手密、安乐割手密、勐根割手密、芒美割手密、贺海割手密、回落割手密、里拉割手密和曼亨割手密,对照材料主要包含了云南、四川、越南、老挝、泰国地区的割手密,其共同特点是均分布在内陆地区;第三类群包括3个材料,分别是海南1号、海南92-2和广东化州割手密,其中不包含新采集的材料。而在相似性系数为0.654处作切割线又能将上述第二类群分为较细的三个亚群。由此可见,新采集的11份割手密资源具有丰富的遗传多样性,与已收集的资源相比,具有一定的差异性。说明依靠云南高山峡谷等立体气候特点,分布着遗传差异显著的割手密无性系。     

间种蔬菜对甘蔗地生态环境和甘蔗生长的影响

甘蔗 (ＳａｃｃｈａｒｕｍｏｆｆｉｃｉｎａｒｕｍＬ .)是广西重要的经济作物 ,种植面积达 4 5 33× 10 6ｈａ[1] ,甘蔗的前期生长比较缓慢 ,行间有比较大的可利用空间 ,到6月下旬～ 7月上旬才进入迅速生长期并逐渐封行。有研究表明 ,在甘蔗园间种玉米对甘蔗的生长发育和产量有一定的影响[5] ,而合理间种豆科作物 (黄豆和花生 )不仅不影响甘蔗的正常生长发育 ,还可以增加甘蔗的产量[4 ,6] 。但间种作物对甘蔗园生态环境的影响未见报道 ,我们于 1997- 1999年在桂西北开展了甘蔗园间种经济效益比较高的蔬菜作物的试验 ,并对间种甘蔗园生态环…     

甘蔗杂交后代蔗汁品质性状的遗传分析

以4×3不完全双列杂交(NCⅡ)选配的12个杂交组合后代为材料,对蔗汁锤度、旋光读数、转光度、蔗汁蔗糖分、视纯度和重力纯度等6个品质性状进行配合力和遗传力分析。结果表明,品质性状的遗传主要由基因加性效应引起,其中蔗汁锤度、旋光读数、转光度、蔗汁蔗糖分主要由父本基因加性效应引起。CP72-330、HoCP93-750、桂糖91-116和粤糖92-1287等4个亲本的品质性状一般配合力(gca)为正值且较大,是配合力较好的高糖亲本。CP72-330×桂糖91-116及HoCP93-750×粤糖92-1287的品质性状特殊配合力(sca)相对效应值均为正值且较大,杂交后代品质性状平均值也较高,为较好的高糖杂交组合。品质性状父本gca方差大于母本gca方差,亲本gca方差为组合sca方差的7-28倍,广义遗传力(hB2)为58%-68%,狭义遗传力(hN2)为56%-60%,属于遗传能力较强的品质性状。     

The Response of Sugarcane (Saccharum officinarum) Cultured Cells to Anoxia and the Selection of a Tolerant Cell Line

The effect of anoxia on the sugarcane (Saccharum officinarum L.) cultured cells was studied in order to elaborate a technique for in vitro selection of cell lines, which would be tolerant to anaerobic stress. Inhibitory and lethal doses of anaerobic incubation were established from the state of the mitochondrial ultrastructure during the anaerobic incubation of cells either with or without exogenous glucose, as well as from the pattern of the post-anaerobic callus growth. An intact state of the mitochondrial ultrastructure and the viability of some cells in the presence of 3% glucose were shown to be maintained for at least 14 days of anaerobic incubation, while the index of post-anaerobic growth decreased by almost 50% even after 72-hour-long anaerobiosis. In the absence of exogenous glucose, a marked destruction of mitochondria and a twofold decrease in the callus growth index were observed as early as after six-hour-long anaerobic stress. A 48-hour-long incubation under these conditions resulted in the maintenance of the intact ultrastructure only in 7–10% of cells, while a 96-hour-long anaerobiosis brought about the complete degradation of the subcellular structure and cell death. A 48-hour-long anaerobiosis without exogenous glucose was chosen for selecting the anoxia-tolerant cell lines. After three cycles of selection, the anoxia tolerance of the selected cell line exceeded the respective index of the initial callus several-fold. In selected line, about 50% of cells retained viability and could resume growth even after 96-hour-long anaerobic incubation. The experimental results obtained were used to determine the possible causes of the heterogeneity of callus cells as regards their anoxia resistance.     

甘蔗过氧化氢酶基因的电子克隆及生物信息学分析

应用电子克隆技术,获得甘蔗中一个过氧化氢酶基因的cDNA序列全长,命名为S-CAT。该基因全长2160bp,包含一个1479bp的开放阅读框,编码492个氨基酸。通过PSORT工具,预测甘蔗S-CAT蛋白存在于过氧化物酶体中。同源比对分析显示,S-CAT基因编码的氨基酸序列与水稻、玉米、高粱、朝鲜碱茅、葡萄等植物中过氧化氢酶基因所编码的氨基酸序列高度同源,同源性分别为97%,97%,95%,91%和92%。研究结果为该基因的实验克隆奠定基础。     

割手密SnRK2基因家族全基因组鉴定及干旱胁迫表达分析

【目的】探究割手密SnRK2基因家族成员在干旱胁迫中的调控机制,为抗旱性甘蔗品种的选育提供侯选基因。【方法】以全基因组数据为基础从割手密中鉴定SnRK2基因,并对其进行生物信息学分析和干旱胁迫下的表达分析。【结果】在割手密基因组中共鉴定出11个SnRK2基因家族成员,命名为SsSnRK2.1-SsSnRK2.11,且这些基因不均匀地分布于8条染色体上。SnRK2蛋白的氨基酸残基数为227～580,分子质量为25 683.53～64 695.8 kD,等电点为4.62～8.94,且均为亲水性蛋白。系统发育树可将其分为3个亚组,且同亚组中的保守基序基本相似,外显子数量以7～9个为主。SsSnRK2基因家族成员的启动子中含有多种激素类和逆境胁迫响应类的作用元件。割手密SsSnRK2基因家族成员的表达具有组织特异性。所有的SsSnRK2基因均能不同程度地响应干旱胁迫。【结论】割手密SnRK2基因家族在响应干旱胁迫过程中发挥重要作用,可为割手密的抗逆性研究提供一定的理论依据。     

甘蔗野生种割手密远缘杂交后代品质性状的遗传研究

利用甘蔗品种Co419与野生种割手密云南75-1-2远缘杂交,ROC25与远缘杂交后代云野02-356进行回交,分别获得F1和BC1群体;利用R软件,分析了2个群体全部真实性后代品质性状的遗传表现.结果表明,杂交后代品质性状广义遗传力高,正态分布特性明显,品质性状间显著正相关;F1含糖量和纤维含量高于商业亲本,但甘蔗蔗糖分、蔗汁糖分、蔗汁锤度和简纯度等性状劣于商业亲本;BC1除含糖量高于双亲外,其他性状介于双亲之间,但主要性状均优于F1,后代性状恢复快.     

云南甘蔗细茎野生种82-114测交后代主要性状遗传分析

利用云南甘蔗细茎野生种82-114(简称:云割82-114)为父本,与甘蔗栽培原种、生产品种和含野生血缘F1等3类亲本进行测交.对后代锤度、茎径、株高、有效茎、蔗产量等5个主要数量性状进行方差、遗传力和配合力分析,结果表明:①组合后代锤度、茎径、株高、有效茎4个性状差异极显著;②组合后代锤度、茎径、株高、有效茎的遗传力表现较突出,均超89.0%;③3类组合中,栽培原种×云割82.114后代有效茎配合力表现明显的正效应,生产品种×云割82-114后代锤度、茎径、株高、蔗产量配合力有比较明显的正效应,舍野生血缘F1×云割82-114后代锤度、茎径、株高、有效茎、蔗产量等5个指标的配合力效应取向不明显.     

斑茅割手密复合体(GXAS07-6-1)及其与甘蔗F_1的GISH分析

斑茅割手密复合体(GXAS07-6-1)是广西蔗茅属斑茅和广西甘蔗属割手密的属间杂种,聚集了双亲的优点。本研究利用基于Alu-like的PCR鉴定方法对GXAS07-6-1及甘蔗与GXAS07-6-1的3份F_1材料进行真实性鉴定,基于基因组原位杂交技术(GISH)对父本GXAS07-6-1及其3份F_1染色体组成及核型进行分析。研究结果表明:3份F_1材料为GXAS07-6-1的真杂种;父本GXAS07-6-1的染色体众数为62条,其中30条来自蔗茅属斑茅,32条来自甘蔗属割手密,核型分类属于1B,其染色体按"n+n"方式传递;GXASF_108-2-17、GXASF_108-2-22、GXASF_108-2-32的染色体数目为78~80条,其中69~71条来自甘蔗属,9~11条来自蔗茅属斑茅,3份F_1的核型分类分别属于2B、1B、1B,染色体传递方式均为"n+n"。父本GXAS07-6-1及3份F_1材料中均未发现有染色体的交换或易位现象。甘蔗与斑割复合体杂交,蔗茅属斑茅染色体在亲子间传递过程存在丢失现象。     

Genetic segregation of microsatellite markers in Saccharum officinarum and S. spontaneum

Genetic mapping techniques can be used to study the interaction between two different genomes after hybridization. This study investigated a Saccharum officinarum (Green German or GG, 2n approximately 11x approximately 110) x S. spontaneum (IND 81-146 or IND, 2n approximately 7x approximately 56) interspecific cross. Segregation of 193 microsatellite (SSR) loci was evaluated in the F(1) progeny of 169 full-sibs of the cross. Following the two-way pseudo-testcross strategy and 'cross pollination' population type, linkage groups (LG) and phases were established for each parent map, using the criteria of LOD score > or = 3.0 and a maximum recombination frequency of 0.35. Of the 193 markers analyzed, 61 were IND-specific, 106 were GG-specific, and 26 were heterozygous in both parents. About 78% of the markers segregated in a Mendelian fashion and 22% were distorted (as evaluated by chi(2)-tests, P < or = 0.01). The GG map included 91 marker loci arranged into 25 LG covering 1180 cM of the officinarum genome. The IND map consisted of 46 marker loci assembled into 10 LG, which spanned 614 cM of the spontaneum genome. A specific chromosome associated with segregation distortion was detected in the female (GG) genome only, probably as a result of double reduction. The segregation patterns of the marker loci indicated a centromere-driven distortion process with the shared allelic markers (as putative centromeres) regulating the placement and association of markers with opposite phase (coupling vs repulsion) and dosage on either side. Although incomplete, the framework maps were informative with respect to segregation distortion, chromosome fusion, rearrangements, and translocations, observed in both parental genomes as a result of their merger.     

Molecular detection of Colletotrichum falcatum causing red rot disease of sugarcane (Saccharum officinarum) using a SCAR marker

Red rot disease of sugarcane caused by Colletotrichum falcatum is one of the most destructive diseases of sugarcane (Saccharum officinarum) worldwide. The pathogen spreads primarily through infected sugarcane setts and hence the use of disease‐free planting materials is essential for preventing disease development in the field. In the present study a polymerase chain reaction (PCR) assay was developed for accurate and sensitive detection of C. falcatum in planting materials. Randomly amplified polymorphic DNA (RAPD) analysis identified a 566 bp PCR fragment that was specific to C. falcatum. The DNA sequence of this fragment was determined and used to design oligonucleotides amplifying a 442 bp sequence characterised amplified region (SCAR). The specificity of the SCAR primers was evaluated using purified DNA from C. falcatum and other Colletotrichum spp. as templates in PCR. The results indicated that the SCAR primers were highly specific to C. falcatum since the 442 bp fragment was amplified only from DNA of isolates and races of C. falcatum but not from any other Colletotrichum spp. tested. The detection sensitivity of C. falcatum was 0.1 ng for genomic DNA of C. falcatum and 5 ng for DNA extracted from infected sugarcane tissue. This new PCR‐based assay is a convenient tool for detection of this important pathogen in seed canes to ensure production of sugarcane.     

Expression of the Grifola frondosa Trehalose Synthase Gene and Improvement of Drought-Tolerance in Sugarcane （Saccharum officinarum L.）

Trehalose Is a nonreduclng dlsaccharlde of glucose that functions as a protectant In the stabilization of blologlcal structures and enhances stress tolerance to abiotic stresses in organisms. We report here the expression of a Grlfola frondosa trehalose synthase （TSase） gene for Improving drought tolerance In sugarcane （Saccharum offlclnarum L.）. The expression of the transgene was under the control of two tandem copies of the CaMV35S promoter and transferred Into sugarcane by Agrobacterium tumefaciens EHA105. The transgenlc plants accumulated high levels of trehalose, up to 8.805-12.863 mg/g fresh weight, whereas It was present at undetectable level in nontransgenlc plants. It has been reported that transgenlc plants transformed with Escherlchla coil TPS （trehalose-6-phosphatesynthase） and/or TPP （trehalose-6-phosphate phosphatase） are severely stunted and have root morphologlc alterations. Interestingly, our transgenlc sugarcane plants had no obvious morphological changes and no growth Inhibition in the field. Trehalose accumulation in 35S-35S：TSase plants resulted In In- creased drought tolerance, as shown by the drought and the drought physiological Indexes, such as the rate of bound water/free water, plasma membrane permeability, malondlaldehyde content, chlorophyll a and b contents, and activity of SOD and POD of the excised leaves. These results suggest that transgenlc plants transformed with the TSase gene can accumulate high levels of trehalose and have enhanced tolerance to drought.     

割手密野生资源抗逆性研究进展

割手密作为现代甘蔗遗传杂交育种史上最为成功的野生亲本,对多种不良环境都具有很强的抗逆性,被公认为是抗逆基因的主要来源。但目前真正被有效利用的割手密抗逆亲本和抗逆基因非常有限,我国自育和引进甘蔗主栽品种的抗逆性仍然比较单一且普遍偏弱,因此加强割手密优良抗逆亲本筛选和抗逆基因挖掘利用研究意义重大。本文综述了不同基因型割手密在非生物逆境(干旱、低温等理化因素)和生物逆境(病虫害侵染)下的抗逆性鉴定及其抗逆基因克隆和功能验证等国内外相关研究进展;并探讨了当前割手密资源抗逆材料筛选和抗逆基因挖掘利用中存在的问题和今后的研究方向,希望为高效利用割手密优异抗逆基因资源开展甘蔗多抗逆性聚合育种提供参考。     

甘蔗MYB2转录因子的电子克隆和生物信息学分析

用电子克隆方法获得甘蔗MYB2基因,采用生物信息学方法,对该基因编码蛋白从氨基酸组成、理化性质、跨膜结构域、疏水性/亲水性、亚细胞定位、高级结构及功能域等方面进行了预测和分析。结果表明:甘蔗MYB2基因全长991bp,包含570bp的ORF,编码189个氨基酸。甘蔗MYB2基因包含有MYB功能域,在序列组成、高级结构及活性位点等方面,与玉米等其它植物的MYB2基因具有高度的相似性。研究结果为该基因的实验克隆奠定基础。     

云南割手密创新种质F_2的主成分聚类分析及其评价

云南割手密是甘蔗育种中比较宝贵的野生种质资源。对68份云南割手密F2创新种质材料的9个主要工农艺性状进行主成分分析,并以前3个主成分为基础,分别作聚类分析和二维排序图。结果表明:68份云南割手密F2创新种质的变异系数为10.57%～35.30%,11月份理论含糖量和单产的变异系数超过30%,说明11月份理论含糖量和单产性状的变异潜力较高。主成分分析结果表明,单茎重产量因子、锤度糖分因子和丛有效茎含糖量这3个主因子提供了原始性状80.8292%的信息;在欧氏距离为10.00处将68份材料分为4类,各个类群具有不同的特点,在甘蔗杂交育种上可以根据不同类群的特点加以利用;二维排序表明云割F211-50、云割F211-56、云割F211-40、云割F211-66、云割F211-77、云割F211-10、云割F211-19、云割F211-85、云割F211-45、云割F211-37、云割F211-25、云割F211-23、云割F211-18等13份材料的3个主成分构成因子协调性最好,可在育种中加以利用。