马铃薯体细胞杂种后代青枯病抗性鉴定及分子标记检测

马铃薯(Solanum tuberosum L.)青枯病是由茄科雷尔氏菌(Ralstonia solanacearum)引起的一种毁灭性的细菌性土传病害。马铃薯青枯病抗性资源主要存在于一些野生种中,体细胞杂交是创制马铃薯青枯病抗性资源的一种有效途径。本研究以具有对青枯病抗性的体细胞杂种与栽培种杂交产生的100个后代为材料,对其进行青枯病抗性评价,旨在筛选可供育种利用的青枯病抗性资源。青枯病抗性鉴定结果表明,在试管苗组培鉴定中100个杂交后代共有6个基因型表型抗青枯病,温室钵栽接种鉴定有8个基因型表现为抗病,在两种接种鉴定中均表现为抗病的有3个基因型(07SF.3-79、07SF.6-8和07SF.6-5)。选用本实验室前期筛选的与青枯病抗性相关的4对SSR标记引物(STI0051、STI0054、STI0056、STI0057),对100个基因型进行分子标记检测,结果显示,有3个标记(STI0051.180、STI0054.180、STI0056.205)可以明确鉴定抗感基因型,它们表现为抗病稳定的3个基因型的标记位点与抗病对照的带型一致,而在感病对照中缺失,与表型鉴定结果吻合,表明筛选的SSR标记可以用于具有S.chacoense遗传背景材料的青枯病抗性辅助选择。     

209份国外春小麦种质对禾谷孢囊线虫的抗性

为了寻找抗禾谷孢囊线虫的春小麦种质资源,于2015-2016年采用两年三个点的田间病圃鉴定和室内二龄幼虫接种鉴定相结合的方法,进行了209份国外春小麦种质对禾谷孢囊线虫的抗性鉴定。结果显示,2015年在大通田间病圃测试中,初筛到抗病材料63份;2016年对63份抗病材料在大通和湟中两个点进行田间验证鉴定,依据Andersen等的标准,得到抗病小麦种质8份;依据 Soriano等的标准,表现抗病的小麦种质11份;按两个标准均表现抗病的材料有7份,为MY00653、MY00865、MY00982、MY001302、MY001307、MY001592和MY002732。室内人工接种鉴定得到抗病春小麦材料4份。田间和室内均表现抗病的材料为MY00653、MY001302、MY001307和MY001592,系禾谷孢囊线虫的潜在抗性资源。     

海岛棉染色体置换系与陆地棉随机交配群体不同株系的黄萎病抗性分析

海岛棉具有纤维品质优良、抗病、抗逆等优点,创制带有海岛棉优异基因的棉花新种质,一直是陆地棉种质创新的一个重要研究方向。海岛棉染色体置换系与陆地棉随机交配群体是具有陆地棉背景而携带众多海岛棉基因的遗传群体,遗传多样性丰富。为筛选抗、耐棉花黄萎病的新种质,采用温室苗期切根接种法和田间病圃成株期鉴定方法,对海岛棉染色体置换系与陆地棉随机交配群体RMBUP-C4S5的148个株系进行黄萎病抗性鉴定。温室苗期切根接种法鉴定结果显示,鉴32、鉴49、鉴59、鉴62、鉴78、鉴83、鉴84和鉴85共8个株系表现为耐黄萎病,相对病情指数（相对病指）为30.0～33.9,占鉴定株系的5.4%;抗黄萎病对照中植棉2号相对病指为34.1,高于上述8个耐黄萎病株系,表现为耐黄萎病。对8个苗期耐病、8个感病的RMBUP-C4S5株系进行田间病圃成株期黄萎病抗性鉴定,结果表明：鉴78、鉴85表现为抗黄萎病,相对病指分别为14.3和13.0,低于当年中植棉2号相对病指22.3;鉴19、鉴49、鉴83、鉴124和鉴143均表现为耐黄萎病。鉴49、鉴78、鉴83和鉴85在苗期耐黄萎病,在成株期表现为抗或耐黄萎病,可作为...     

优质两系杂交稻组合两优2163稻瘟病抗性研究初报

用属5个不同生理小种的11个广谱致病单孢菌株和广谱致病多孢菌株对两优2163进行苗期人工接种鉴定,同时在福建省10个稻瘟病鉴定圃进行田问自然诱发鉴定。鉴定结果苗期人工接菌鉴定1998年表现感(S),1999年表现中抗(MR),田间病圃自然诱发鉴定1998年叶癌表现感(S),穗颈瘟表现中抗(MR),1999年叶瘟表现为感(S),穗颈瘟表现为中感(MS),两年鉴定综合评价为感稻瘟病(S),生产上推广时要注意稻瘟病的防治。     

花生种质资源抗花生青枯病鉴定

通过对3381份(国内2257,国外1124份)花生种质资源的田间自然病圃和人工接种鉴定,共获得高抗花生青枯病的种质资源55份,中抗43份。其中国内资源高抗的50份,中抗的39份,国外资源高抗的5份,中抗4份。不同类型的资源以及同一类型不同地区来源的资源其抗性表现差异极大。并筛选出一批经济性状优良的高抗种质。     

东北春玉米区玉米栽培品种对大斑病抗性水平研究

通过人工接种,利用分离纯化的玉米大斑病强毒力菌株和混合菌株对2008～2009年从东北地区收集的269份玉米品种进行温室和大田玉米大斑病抗性水平鉴定。结果表明,温室鉴定条件下这些栽培品种对玉米大斑病抗性水平较低,田间人工接种和自然发病处理整体表现抗玉米大斑病水平较高。田间人工接种处理高感病品种(HS)占0.37%,感病(S)品种占4.83%,中抗(MR)品种占31.60%,抗病(R)品种占55.02%,高抗(HR)品种占7.80%;温室人工接种处理高感品种占64.31%,感病品种占21.56%,中抗品种占10.40%,抗病品种仅占3.72%;田间自然发病处理高感病品种为0,感病品种占1.11%,中抗品种占10.40%,抗病品种占22.68%,高抗品种占65.80%。     

小麦品种对禾谷孢囊线虫（Heterodera avenae） 江苏沛县群体的抗性鉴定

为了筛选对禾谷孢囊线虫病(Cereal cyst nernatode,CCN)具有抗性的小麦品种,采用室内二龄幼虫接种法、网室卵接种法和田间病圃法测定了收集自5个省份的40个小麦品种对小麦孢囊线虫江苏沛县群体的抗性。室内二龄幼虫接种测定结果表明,供试材料中无免疫品种,华麦1号、温粮58和豫麦66-18系表现为高抗,扬麦13和徐麦7086表现为中抗,烟农5158表现最感病,平均单株白雌虫数达104.7个;田间病圃测定表明,上述5个品种仅华麦1号表现为抗病。网室卵接种法测定结果与室内二龄幼虫接种法及田间病圃法测定结果差异较大,显示该方法不适用小麦品种的CCN抗性鉴定。     

马铃薯抗青枯病和低温糖化体细胞杂种的鉴定

ACC-1-1和ACC-1-2再生株系由来自马铃薯二倍体栽培种(Solanum tuberosum)品系AC142-01(2n=2x=24)和二倍体野生种S.chacoense的一个无性系C9701(2n=2x=24)经原生质体融合获得,经鉴定为体细胞杂种。流式细胞仪分析和染色体计数显示,ACC-1-1为混倍体,ACC-1-2为八倍体。两个株系均能正常开花但花粉活力较低。花粉母细胞减数分裂观察显示,两个系在减数分裂各个时期均出现广泛的异常染色体行为,可能是花粉活力低的原因。青枯病接种鉴定表明,ACC-1-2对青枯病表现为高抗,而ACC-1-1表现为中感,同时这两个株系均具有"低温糖化"抗性,证明体细胞融合加上适当选择可以有效聚合有性杂交不亲和双亲的优良性状。     

小麦种质P10078的成株期条锈病抗性特征及遗传规律

为明确小麦抗病种质P10078的抗条锈病特征及抗病遗传规律,利用4个小麦条锈菌小种CYR32、CYR33、V26/CM42和Sull-7对P10078进行了苗期分小种鉴定,并分别在陕西杨凌人工接种病圃和甘肃天水自然诱发病圃连续多年对P10078与感病品种铭贤169及其杂交后代(F_1、F_2、F_(2∶3))进行成株期抗条锈病鉴定。结果发现,P10078苗期对CYR32、V26/CM42表现为感病(IT:7~8),对V26/CM42和Sull-7表现为抗病(IT:2),成株期对混合小种表现为高度抗病(IT:1,S:1%),表明P10078为包含全生育期抗病基因的成株期抗病品种;F_2抗感分离比为3∶1,F_(2∶3)抗感分离比为1∶2∶1,表明P10078成株期抗性由一对显性主效基因控制。P10078所含抗条锈基因为未知抗性基因。     

大豆抗胞囊线虫4号生理小种SSR标记筛选及优异种质鉴定

大豆抗胞囊线虫的表型鉴定工作量较大,鉴定结果易受环境影响,是抗源筛选和抗病品种选育的限制因素之一。不受时间、环境限制的分子标记鉴定为抗病鉴定提供了一种高效快捷准确简单的鉴定方法。为筛选可用于抗大豆胞囊线虫4号生理小种分子标记引物,本研究采用基因型鉴定和人工接种鉴定相结合的方法鉴定193份大豆资源的大豆胞囊线豆抗性。以部分高抗大豆资源和高感资源为材料,对1 000对SSR引物进行初筛,用初筛获得的引物扩增193份抗、感大豆资源,筛选抗胞囊线虫4号生理小种(SCN 4)SSR标记引物,用于大豆抗胞囊线虫4号生理小种分子标记辅助鉴定。结果表明:筛选出4个与大豆胞囊线虫4号生理小种相关的SSR标记,分别为Satt400、Satt680、Satt533和Satt504。Satt400、Satt680、Satt533和Satt504对感病资源的检出率均为100%,对抗病资源的检出率分别为70.58%、63.15%、92.3%和57.14%。结合人工接种鉴定结果显示,4个标记组合鉴定可提高对胞囊线虫4号生理小种抗性的选择效率,达100%。经人工接种鉴定,从193份资源中筛选出12份抗病资源,其中10份资源为高抗,2份资源为中抗。这12份抗源的4个SSR标记带型均为抗病带型,其余感病和高感资源的4个SSR标记带型则至少有一个为感病带型。因此,利用分子标记评价抗感特性的标准为被1个或1个以上的感病带型检出的资源为感病资源,当4个标记均为抗病带型,该资源为抗病资源。利用该引物组合对山西省种质库提供的100份资源进行鉴定。筛选出4份抗源,人工接种鉴定结果表明,黑豆、小颗黑和晋1265(茶)3份资源抗性级别为高抗,大黑豆抗性级别为中抗。因此,该引物组合可有效辅助鉴定大豆对胞囊线虫4号生理小种的抗性。     

利用体细胞杂交获取马铃薯软腐病的抗性

对马铃薯四倍体栽培种和二倍体野生种Solanumbrevidens的体细胞杂种通过叶片离体培养获得的四倍体植株,以及用四倍体栽培种进行回交获得的五倍体植株的块茎对软腐病的抗性进行了测定。结果表明,由体细胞杂种通过叶片组织离体培养再生植株中,有一个株系SC107对软腐病菌Erwiniacarotovora具有较强的抗性。在用不抗软腐病的马铃薯栽培种对体细胞杂种进行回交获得的杂种后代中,大部分株系对软腐病菌具有高水平的抗性,从而说明Solanumbrevidens对软腐病的抗性基因已转移到马铃薯栽培种。     

Bacterial wilt resistance in the progenies of the fusion hybrids between haploid of potato andSolanum commersonii

Bacterial wilt of potatoes, which used to be a widespread disease in tropics and subtropics, has become a threat to potato production in temperate region. The diploid species Solanum commersonii has several desirable characteristics including cold tolerance and resistance to several diseases. Selected somatic hybrids between S. tuberosum dihaploid and bacterial-wilt-resistant S. commersonii clones were cross pollinated with S. tuberosum cultivars for further selection of bacterial wilt resistance. The chromosome numbers of the fusion parents were confirmed as 24, and the three fusion hybrids crossed were all tetraploids. The chromosome number of 11 backcross 1 progenies (BC1) was 48 and that of the other six was close to the tetraploid number. Backcross 2 progenies (BC2) were obtained from only three of the 44 BC1 clones crossed. The S. commersonii parent clone, LZ3.2, was the most resistant to bacterial wilt among wild species clones tested. The first sexual progenies segregated for resistance, with one clone highly susceptible and four clones highly resistant. Three highly resistant BC1 clones, CT02-4, CT08-4, and CT10b-4, were backcrossed to cultivars. Two cross combinations produced mostly susceptible BC2 progenies; however seven clones were resistant or highly resistant for both race 1 and race 3. The highly resistant three clones, CT204-3, CT206-9 and CT206-10, were selected for the further testing as cultivars or breeding materials.     

马铃薯抗青枯病体细胞变异体离体筛选研究初报

本试验证明了青枯菌活菌作为马铃薯细胞变异体离体筛选的选择压力是适宜的。明确了活菌压力的最适浓度、愈伤组织接种的最适龄期和接种的方法。愈伤组织培养皿内继代培养重复接种筛选是快速而有效的筛选方法。从近2万块马铃薯叶圆片愈伤组织中,经重复接种筛选获得了6块抗菌愈伤组织(比率为0.03％左右)。并经过分化诱导培养获得了再生苗。初步的抗性鉴定结果表明,再生苗抗1(PSS—1)比母体(米拉脱毒试管苗)的抗病性有显著提高。     

Soft rot and blackleg reactions in diploid potato hybrids inoculated withErwinia spp.

From 1993 to 1996 three groups of potato genotypes were evaluated for resistance toErwinia spp.: (1) 31 interspecific diploid hybrids (28 resistant and three susceptible), (2) five hexaploid or pentaploid somatic hybrids ofSolanum tuberosum (tbr) ×S. brevidens (brd), and (3) eight cultivars. Two evaluation methods were applied: tuber point inoculation with eitherErwinia carotovora ssp.atroseptica (Eca) orE. chrysanthemi (Ech) to test tuber soft rot resistance and stem base inoculation with Eca to test blackleg resistance. Some resistant diploid hybrids and somatic hybrids oftbr × brd were superior to cultivars for both tuber and stem resistance. Tuber and stem resistance toErwinia spp. in the most resistant diploid hybrids were equal to the highly resistant somatic hybrids oftbr xbrd. Tuber resistance to Eca was highly correlated to tuber resistance to Ech (r=0.815***). In two years of evaluation for stem resistance, three diploid hybrids and a derivative of one of the somatic hybrids (USA M 264) failed to develop symptoms of blackleg following inoculation with Eca, Analyses of variance for tuber and stem resistance indicated significant effects of genotype, year and genotype × year interaction. A positive relationship between tuber and stem resistance toErwinia spp. has been found, however the genetic control of resistance in tuber and stem is partially independent. In the case of Eca the correlation coefficient was r = 0.725***. Therefore it should be possible to obtain resistant genotypes to both blackleg and tuber soft rot. Several resistant diploid hybrids were selected from among those tested, which also have several other characters desirable for potato breeders.     

An <Emphasis Type="Italic">In Vitro</Emphasis> Assay Method for Resistance to Bacterial Wilt (<Emphasis Type="Italic">Ralstonia solanacearum</Emphasis>) in Potato

To develop an in vitro assay method for bacterial wilt resistance in potato, resistant and susceptible standard genotypes were grown in vitro, and different conditions of inoculation with Ralstonia solanacearum phylotype I/biovar 4 were examined. The optimal condition was the inoculation of 6–8 leaf stage plants with a bacterial concentration of 10² CFU ml^?1 and an incubation temperature of 28 °C. Evaluation of stem wilting was more reliable than that of leaf wilting. Using this method, nine genotypes with different resistance levels in the field were evaluated. Lower disease indices were obtained for genotypes with high resistance levels in the field, suggesting that this assay is useful for evaluating bacterial wilt resistance in a controlled environment.     

Comparison of the effectiveness of different methods of screening for bacterial soft rot resistance of potato tubers

Tubers from eight potato cultivars (cvs) grown at two different locations in Wisconsin were tested for bacterial soft rot resistance using different inoculation techniques. The procedures included 1) point inoculations of tubers with different inoculum levels followed by incubation in ambient or low oxygen condtions, 2) inoculation of mechanically bruised tubers followed by incubation in a mist chamber, and 3) a standard slice inoculation method. The point titration test followed by incubation in dew chamber and the mist chamber-bruise test showed similar patterns of resistance for cultivars that were used in these experiments. These two methods are considered to be effective for screening potato tubers for bacterial soft rot resistance. Point titration methods are very useful if only limited numbers of tubers are available. The mist chamber-bruise test is simpler than the other procedures; however, to obtain reproducible results large numbers of tubers are required. Because of the great variability of the results obtained in inoculation of slices, the reliability of this approach can be questioned as a standardized method for evaluation of resistance. Tubers of somatic hybrids ofS. brevidens andS. tuberosum and their sexual progeny were significantly more resistant to bacterial soft rot than tubers of moderately resistant cultivars when evaluated by each of the assay procedures.     

Breeding Potential of Potato Somatic Hybrids: Evaluations for Adaptability,Tuber Traits,Late Blight Resistance,Keeping Quality and Backcross (BC<Subscript>1</Subscript>) Progenies

Ten tetraploid interspecific potato somatic hybrids developed earlier through protoplast fusion between the dihaploid Solanum tuberosum L. ‘C-13’ and the diploid wild species Solanum pinnatisectum Dun. were used in this study. Somatic hybrids and standard control cultivars were evaluated for adaptability, tuber traits, late blight resistance and keeping quality attributes (dormancy, sprouting, weight loss and appearance after 75 days of storage) during two successive winter crop seasons (short-days) in the sub-tropical plains of India, where nearly 90% of India’s potatoes are grown. Somatic hybrids showed medium to good plant vigour and late to very late plant maturity. Though none of the somatic hybrids produced equal or higher tuber yield than the control cultivars, except clone P7, all other somatic hybrids produced significantly higher marketable and total tuber yield after a 90-day growth than the parent C-13. Most of the somatic hybrids possessed higher tuber dry matter concentration, resistance to late blight and better keeping quality attributes than the control cultivars. Notably, the tubers of somatic hybrids showed a tendency for colour change from white to purple on exposure to sunlight. Breeding potential of somatic hybrids was further assessed by crossing them with commonly grown local potato cultivars for the development of backcross (BC₁) progenies. In the hybridization programme, berries were formed while somatic hybrids were used either as male or female parents with common cultivars. Our results revealed that the use of bulk pollen of somatic hybrids not only resulted in higher pollination success but also helped to achieve higher numbers of true potato seeds (TPS) per berry. Despite the formation of berries, TPS was absent in 22 crosses using somatic hybrids as a female or male parent which may be due to ovule or embryo abortions. Segregating BC₁ generations were evaluated under the same sub-tropical conditions for further exploitation in potato breeding. Based on our study, the somatic hybrids P4, P8 and P10 can be utilized for the introgression of important characters such as high tuber dry matter concontration, resistance to late blight and excellent keeping quality attributes into the cultivated potato via conventional breeding methods for cultivar development in the sub-tropical plains of India.     

Transfer of PLRV resistance fromSolanum verrucosum Schlechdt to potato (S. tuberosum L.) by protoplast electrofusion

Summary Somatic fusions between an accession of the diploid wild speciesSolanum verrucosum and a dihaploid S.tuberosum genotype were produced in order to incorporate resistance to potato leafroll virus (PLRV). In total 15 somatic hybrids out of 16 regenerants were obtained. Identification of hybrids was based on additive RAPD patterns, general morphological characteristics, chromosome numbers and chloroplast counts in stomata guard cells. A field trial was performed with the hybrids, their two parents and the control cultivar Kennebec to assess field performance and phenotypic variability. Yield parameters varied considerably among somatic hybrids. Some of the hybrids gave significantly higher yields, tuber numbers and tuber weights than both parents. Pollen fertility of hybrids ranged from 19 to 77%. Twelve hybrids were found to be resistant to PLRV.     

Interspecific somatic hybridization betweenSolanum tuberosum L. andS. bulbocastanum dun. as a means of transferring nematode resistance

Interspecific somatic hybrids were produced between tetraploidSolanum tuberosum and a nematode-resistant accession of the diploid speciesSolanum bulbocastanum by protoplast fusion. Hybrid cells were selected using dual fluorescent labeling of protoplast preparations prior to fusion. Hybridity of regenerated plants was confirmed with a combination of morphological assessment, chromosome counting and isozyme analysis. Somatic hybrids had the same level of resistance to infection by race 1 of the nematodeMeloidogyne chitwoodi as theS. bulbocastanum parent used in the fusion. Some of the somatic hybrids were fertile as females when crossed with tetraploidS. tuberosum breeding lines. Thus, these hybrids can be used in a potato improvement program to incorporate a valuable pest resistance