摘要
多花黑麦草(Lolium multiflorum Lam．)为黑麦草属中最具利用价值的牧草之一，也是我国南方及其它冷凉地区栽培利用面积最广的冷季型牧草。病害是影响多花黑麦草饲草产量和品质的重要因素，多花黑麦草灰叶斑病是由病原真菌Pyricularia sp.引起的一种严重病害。遗传连锁图的构建及基于基因组DNA分子标记的开发将有助于多花黑麦草的遗传及育种研究。本研究以多花黑麦草为材料，利用简并引物PCR扩增法进行多花黑麦草抗病基因类似物(RGA)的克隆并根据获得的RGA序列设计STS引物；以多花黑麦草细胞质雄性不育分离群体(CMS群体)为材料，以SSR、AFLP、EST-CAPS和RGA-CAPS等分子标记构建多花黑麦草遗传连锁图；以多花黑麦草灰叶斑病抗病植株和感病植株杂交构建F_1抗、感病分离群体，分别采用RGA-CAPS、EST-CAPS以及AFLP等分子标记方法进行抗病基因的分子标记和分析，主要结果如下：
1．多花黑麦草RGA的克隆
利用简并引物和巢式PCR扩增克隆法从多花黑麦草基因组中获得24000个300-500bp的DNA片段，采用单通道法对所得DNA序列进行测序，对所得序列进行公共数据库的搜索。结果表明，9344个克隆的DNA序列同已知的抗病基因或从其它植物中获得的RGA序列同源性较高，根据所得克隆的核苷酸序列的相似性，通过聚类分析将其分为185个簇，每簇中选取一个具代表性克隆作为多花黑麦草的抗病基因类似物，对185个抗病基因类似物序列进行开放阅读框的检查，舍弃无效序列，最后对115个被选择的克隆序列进行STS引物设计，成功地设计RGA-STS引物113对。
2．多花黑麦草遗传连锁图的构建
利用多花黑麦草假测交细胞质雄性不育(CMS)分离群体(124个个体)，以来自父本和母本的分离位点采用SSR、AFLP、EST-CAPS和RGA-CAPS等标记方法进行多态性的选择和并对父、母本分别构建遗传连锁图。母本连锁图包含362个标记位点，分别包含SSR标记240个、AFLP标记84个、EST-CAPS标记24个和RGA-CAPS标记14个，整个连锁图由LG1—LG7共7个连锁群组成，覆盖全长776.4cM，7个连锁群中最长的为LG3，135cM，最短的为LG5，85.8cM。相邻标记之间的平均距离为2.14cM。以来自父本的分离位点进行作图所得的LG1—LG7共7个连锁群，包含376个标记，其中SSR标记252个、AFLP标记82个、EST-CAPS标记29个和RGA-CAPS标记13个，连锁图全长710.0cM，7个连锁群中最长的为LG6，117.3cM，最短的为LG7，为80.5cM，相邻标记之间的平均距离为1.89cM。同已报道的多年生黑麦草遗传连锁图相比，本研究中所构建的遗传连锁图具有标记密度高、标记分布均匀和连锁图长度中等等特点。
3．黑麦草RGA在基因组中的分布
应用由RGA序列设计的113对RGA-STS引物进行CMS群体中各个体基因组DNA的PCR扩增，扩增产物分别以识别4到6对碱基的限制性内切酶MseⅠ、AluⅠ，HpyCH4Ⅳ、ScrFⅠ、Sau96Ⅰ、Epy188Ⅲ、DdeⅠ、NlaⅣ、HpyCH4Ⅲ，Fnu4HⅠ、Hpy188Ⅰ，AfaⅠ、MspⅠ、HaeⅢ、HhaⅠ、MboⅠ、HindⅢ、和HinfⅠ共18种进行酶切，酶切产物经2.0％琼脂糖凝胶电脉、显像和多态性检测，共检测到25对引物及相应的限制性内切酶酶切后产生多态性。所检测到的多态性位点经遗传作图，23个标记被标于多花黑麦草遗传连锁图中。结果表明多花黑麦草RGA分布于整个基因组中，但局部有RGA的聚集，其中标记RG124E11-2、RG001E01-2和RG088C11-1聚集于LG2，之间的遗传距离为8.5cM，标记RG192G09-2、RG073A04-3和RG119F12-1聚集于LG7，之间的遗传距离为10.2 cM，标记RG021F04-2和RG227D4-1聚集于LG3，标记间距离仅为2.0cM。RGA-CAPS标记聚集的位点可能存在多花黑麦草抗病基因。
4．多花黑麦草抗灰叶斑病的分子标记
以多花黑麦草灰叶斑病抗病个体(Sachiaoba)和敏感个体(Minamiaoba)杂交构建F_1分离群体，F_1群体的致病菌接鉴定结果显示多花黑麦草抗灰叶斑病由单个主效基因(LmPi1)控制。采用分群分离分析(BSA)法进行多花黑麦草抗灰叶斑病的AFLP和EST-CAPS以及RGA-CAPS标记筛选，筛选到27个标记同多花黑麦草抗灰斑病基因LmPi1紧密连锁，其中AFLP标记25个、EST-CAPS标记1个和RGA-CAPS标记1个。经连锁分析，结果表明，27个标记构成一个紧密连锁的连锁群，标记间最大遗传距离19.6 cM。其中12个标记包括1个EST-CAPS标记和11个聚集于相同的位点，经QTL分析，该连锁区间支持1-LOD，进一步证实本研究中多花黑麦草抗灰叶斑病由单基因所控制。EST-CAPS标记p56的限制性酶切片段同多花黑麦草LmPi1位点的抗病等位基因一致。参考对照群体的遗传连锁图，抗病基因LmPi1位于多花黑麦草的第五连锁群(LG5)。标记p56试用于多花黑麦草抗灰叶斑病抗病和感病品种的基因型分析结果表明，标记p56对于感病种质中引入抗病基因LmPi1以提高其抗病性具有重要作用。
多花黑麦草抗病基因类似物的克隆以及遗传连锁图的构建将为多花黑麦草抗病基因的分子标记、抗病基因的图位克隆以及抗病育种提供了方便。
Italian ryegrass (Lolium multiflorum Lam) is one of the most important temperate forage grasses and is widely cultivated in temperate areas of the world and also in the southern of China. Disease is one of the main factors, which affects its forage yield and quality of Italian ryegrass. Ryegrass blast, also called gray leaf spot, is caused by the fungus Pyricularia sp. It is one of the most serious diseases for Italian ryegrass in China and other temperate regions. With construction of genetic linkage map and development of molecular marker based on DNA, it would facilitate genetic research and breeding of Italian ryegrass. The present study was designed according to the NBS-LRR motif of disease resistance gene by isolating the resistance gene analogs from Italian ryegrass using PCR amplification, multiple combinations of degenerate primers and construct a high-density molecular linkage map of Italian ryegrass, one pseudo-testcross full-sib F_1 population consisting of 124 individuals was used to analyze four types of marker: (1) amplified fragment length polymorphism (AFLP) markers, (2)simple sequence repeat (SSR) markers, (3)expressed sequence tag cleaved amplification polymorphism sequence (EST-CAPS) markers and (4)resistance gene analog CAPS (RGA-CAPS) markers. An F_1 population, consisted 162 individuals, composed from a cross between resistant and susceptible cultivars, was used to screen closely linked markers to resistance gene of gray leaf spot disease by AFLP, EST-CAPS and RGA-CAPS. The main results are as follows:1. RGA isolated from the genome of Italian ryegrassThe large-scale cloning of resistance gene analogs (RGAs) was determined in Italian ryegrass. The degenerate primer designed from conserved motifs of known plant resistance gene products were used to amplify genomic DNA sequences by nested PCR amplification. Total 24000 PCR clones were selected and subjected to one pass sequencing for the sequence similarity search with public database. As a result, 9354 clones showed significantly similar levels of identity compared with known resistance genes or RGAs derived from other species. All these clones were clustered and grouped into 185 clusters based on the nucleotide sequence similarity. The representative clone was selected from each cluster and then was defined as Italian ryegrass RGAs. 150 representative clones, which contains continuous open reading frame, they were selected to covert RGAs to STS markers, 113 specific sequence tagged site (STS) primer sets were designed successfully from these RGA sequences.
2. Genetic linkage map of Italian ryegrass
One pseudo-testcross full-sib F_1 population generated for the genetic analysis of cytoplasmic male sterility (CMS) analysis, consisting of 124 individuals, was used to construct a high-density linkage map of Italian ryegrass. Male and female molecular-marker linkage maps were developed using SSR, AFLP, EST-CAPS, RGA-CAPS markers, respectively. In the linkage map of female parent, a total of 362 marker loci, which was consisted of 240 SSR markers, 84 AFLP markers, 24 EST-CAPS markers and 14 RGA-CAPS markers, they were divided into seven linkage groups and covered a total length of 776.4cM. The size of the linkage groups ranged from 85.8 to 135.0 cM with an average of 2.14 cM among markers. The male linkage map was consisted of 376 marker loci, 252 SSR markers, 82 AFLP markers, 29 EST-CAPS markers and 13 RGA-CAPS markers among seven groups, which covered the map distance 710.0 cM. The longest and shortest linkage groups were LG6 and LG7, 113.7 cM and 80.5 cM, respectivcly. The average distance is 1.89 cM among markers. These linkage maps were compared with those constructed from perennial ryegrass, the map constructed in this research showed very high-density of markers, moderately in length, and markers distributed evenly.
3. Distribution of RGA in the genome of Italian ryegrass
CMS F_1 population (consisting 124 individuals) and 113 pairs of RGA primers were used for RGA mapping. Eighty pairs of RGA primers were amplified with one clear anticipated STS (300-600bp), and then were selected. Using of the 80 pairs of primers to amplify the genomic DNA of two parents and individuals of F_1 population, PCR products digested with endonucleases AluⅠ, AfaⅠ, MboⅠ, MspⅠ, MseⅠ, HinfⅠ, Sau96Ⅰ, HindⅢ, SerFⅠ, HpyCH4Ⅲ, HpyCH4Ⅳ, NlaⅣ, HhaⅠ, Hpy188Ⅰ, Hpy188Ⅲ, HaeⅢ; DdeⅠand Fnu4HⅠ, respectively. Twenty-five polymorphisms were detected and 23 RGA-CAPS markers were mapped to all the seven genetic linkage groups of Italian ryegrass. RGA-CAPS markers distribute all the genome, however, clustered in some loci. Two threemarkers and one two-markers were clustered on the genetic map. Marker RG124E11-2, RG001E01-2 and RG088C11-1 clustered on LG2 within 8.5 cM, marker RG192G09-2, RG073A04-3 and RG119F 12-1 clustered on LG7 within 10.2 cM, marker RG021 F04-2 and RG227D4-1 were mapped on LG3 within 2.0 cM. The region clustered RGA-CAPS markers maybe existed disease resistance locus or loci.
4. Screening molecular markers linked to resistance gene of gray leaf spot in Italian ryegrass
An F: population with a cross species from a resistant (Sachiaoba) and a susceptible (Minamiaoba) cultivar was analyzed. The disease severity distribution in the F_1 population suggested that the disease resistance was controlled by a major gene (LmPil). Analysis of amplified fragment length polymorphisms (AFLP) with bulked segregant analysis identified several markers tightly linked to LmPil. To identify other markers linked to LmPil, expressed sequence tag cleaved amplified polymorphic sequence (EST-CAPS) markers and resistance gene analog CAPS (RGA-CAPS) markers were also screened and analyzed. One EST-CAPS marker, p56, and one EST-CAPS marker, RG036G04-1, were also found to be closely linked to LmPil locus. Twenty-seven markers were mapped to a closely linked group with 19.6 cM in length. Furthermore, twelve markers including one ESI-CAPS marker, p56, and other eleven AFLP markers clustered in one locus. QTL analysis indicated that the region clustered markers supported 1-LOD interval, and the result also indicated one major gene controlled the disease resistance. The restriction pattern of p56 amplification showed a unique fragment corresponding to the resistant allele at the LmPi1 locus. A nkage map constructed from the reference population showed that the LmPi1 locus was located in linkage group 5 (LG5) of Italian rycgrass. Genotype results obtained from resistant and susceptible cultivars indicate that the marker p56 is useful for introduction of the LmPi1 gene into susceptible germplasm in order to develop Italian ryegrass cultivars with enhanced resistance to Italian ryegrass gray leaf spot disease.
The cloning of RGAs, construction of high density genetic linkage map and development of molecular markers closely linked to gray leaf spot disease resistance gene will provide for gene targeting, quantitative trait loci mapping, and marker-assisted selection in Italian ryegrass.
引文
[1]杨中艺，辛国荣．“黑麦草—水稻”草田轮作系统应用效益初探(案例研究)[J]．草业科学，1997，(6)：35-38．
[2]潘水年．黑麦草优质高产栽培技术[J]．中国草地，1996，(1)：24-26．
[3]陈三有．从广东草业生产模式看牧草在可持续发展中的战略地位[J]．草业科学，2000，(2)：75-78．
[4]唐积超．冬闲田种黑麦草效益高[J]．草与畜，1998(2)：37-38．
[5]王进波，齐莉莉．优质饲草—黑麦草的开发与利用[J]．饲科研究，2000，(10)：27-28．
[6]沈益新，杨志刚，刘信宝．凋萎和添加有机酸对多花黑麦草青贮品质的影响[J]．江苏农业学报．2004，20(2)：95-99．
[7]Clarke BB, Vaiciunas SS. Best management practices for the control of gray leaf spot. Phytopathology. 2001,91:148.
[8]Bain DC, Patel BM, Patel MV. Blast of ryegrass in Mississippi[J]. Plant Dis. Rep. 1972, 56:210.
[9]Carver RB, Rush M, Lindberg GD. An epiphytotic of ryegrass blast in Louisiana[J]. Plant Dis. Rep. 1972, 56:157-159.
[10]Sumida Y, Fukubara H, Inoue T, Kajihara H, Mizuno K, Fujiwara T. A study of the occurrence of dieback on Italian ryegrass (Lolium multiflorum) caused by Pyricularia sp.Investigation of facts about occurrence and development of a new resistance test method on breeding (in Japanese)[J]. Bull Yamaguchi Agric Exp Stn.2003, 54:29-36.
[11]Landschoot PJ, Hoyland BF. Gray leaf spot of perennial ryegrass turf in Pennsylvania [J]. Plant Dis. 1992, 76:1280-1282.
[12]Trevathan LE, Moss MA, Blasingame D. Ryegrass Blast [J]. Plant Dis. 1994, 78:113-117.
[13]Viji G, Wu B, Kang S, Uddin W, Huff DR. Pyricularia grisea causing gray leaf spot of perennial ryegrass turf: population structure and host specificity. Plant Dis [J]. 2001, 85:235-240.
[14] Vincelli, P. Gray leaf spot: An emerging disease of perennial ryegrass [J]. Turfgrass Trends 1999, 7:1-8.
[15] Mizuno K, Yokohata Y, Oda T, Fujiwara T, Hayashi K, Ozaki R, Kobashi K, Ashizawa H, Ushimi T. Breeding of a new variety 'Sachiaoba' in Italian ryegrass (Lolium multiflorum L.) and its characteristics (in Japanese)[J]. Bull Yamaguchi Agric Expt Stn. 2003, 54:11-24.
[16] Uddin W, Soika MD, Moorman FE, Viji G. A serious outbreak of blast disease (gray leaf spot) of perennial ryegrass in golf course fairways in Pennsylvania[J]. Plant Dis. 1999, 83:783.
[17] Giesler LJ, Yuen GY, Horst GL. Canopy microenvironments and applied bacteria population dynamics in shaded tall fescue [J]. Crop Sci. 2000,40:1325-1332.
[18] Williams DW, Burrus PB, Vincelli P. Severity of gray leaf spot in perennial ryegrass as influenced by mowing height and nitrogen level [J]. Crop Sci. 2001, 41:1207-1211.
[19] Trevalan LE. Response of ryegrass introductions to artificial indoculation with Pyricularia grisea under greenhouse conditions. Plant Dis. 1982, 66:696-697.
[20] Stacy AB, Christine K, Bruce BC, William A. Meyer. Breeding perennial ryegrass for resistance to gray leaf spot [J]. Crop Sci. 2004, 44: 575-580.
[21] Curley J, Sim SC, Jung G, Leong S, Warnke S, Barker RE. QTL mapping of ryegrass blast resistance in ryegrass, and synteny-based comparison with rice blast resistance genes in rice[M]. In: Hopkins A, Wang ZY, Mian R, Sledge M, Barkers RE(eds) Molecular breeding of forage and turf. Kluwer, Dordrecht, 2004, pp 37-46.
[22] Flor HH. Current status of the gene-for-gene concept [J]. Annu Rev Phytopathol. 1971, 9:275-296.
[23] Lili M, Justin DF, Robert LB, Dikram SG, and John PF. Physical and Genetic Mapping of Wheat Kinase Analogs and NBS-LRR Resistance Gene Analogs [J]. Crop Sci, 2003, 43: 660-670.
[24] Dixon MS, Jones DA, Keddie JS, et al. The tomato Cf-2 disease resistance locus comprises two functional genes encoding leucine rich repeat proteins [J]. Cell, 1996, 84: 451-459.
[25] Song WY, Wang GL, Chen LL, et al. A receptor kinase-link protein encoded by the rice disease resistance gene, Xa21 [J]. Sciences, 1995,270:1804-1806.
[26] Martin GB, Brommmonschenkel SH, Chunwongse J, et al. Map-based cloning of a protein kinase gene conferring disease resistance in tomato [J]. Science, 1993,262:1432-1436.
[27] Joosten M, De Wit P. The tomato-Cladosporiom fulcum interaction: a versatile experimental system to study plant-pathogen interactions [J]. Annu Rev. of Phytopatholigy, 1999,37:335-367.
[28] Whitham S, Dinesh-Kumar SP, Choi D, et al. The product of the tobacco mosaic virus resistance gene N: Similarity to toll and the interleukin-1 receptor [J]. Cell, 1994,78:1011-1015.
[29] Johal G S, Briggs S P. Reductase activity encoded by the Hm 1 disease resistance gene in maize [J]. Science, 1992,258:985-987.
[30] Buschges R, Hollricher K, Panstruga R, et al. The barley mlo gene: a novel control element of plant pathogen resistance [J]. Cell, 1997, 88(5):695-705.
[31] 朱立煌，徐吉臣，陈英等．用分子标记定位一个未知的抗稻瘟病基因[J]．中国科学(B辑)，1994，24：1084-1052．
[32] Young ND, Zamir D, Ganal MW et al. Using of isogenic Lines, and simultaneous probing to identify DNA markers tightly linked to the Tm-2a gene in tomato [J]. Genetics, 1988,120:579-585.
[33] Penner GA. Identification of RAPD markers linked to a gene govering cadmiun uptake in durum wheat [J]. Genome. 1995,38:543-547.
[34] Fehrmann H, Dimond AE. Peroxidase activity and phytophthora resistance in different organs of the potato plant [J]. Phytopath. 1987, 57:69-72
[35] Ye YS, Pan SQ, Kuc S. Association of pathogenesis-related proteins and activities of peroxidase β-1,3-glucanase and chitinase with systemic induced resistance to blue mould of tobacco but not to systemic tobacco mosaic virus [J]. Physiol. Mol. Plant Pathol., 1990, 36:523-528.
[35] Schlumbaum A, Mauch F, Vogeli U et al. Plant chitinases are potential inhibitors of fungal growth [J]. Nature, 1986, 324:365-369.
[36] Shimoni M, Bar-Zur A, Reuveni R. The relation between isozymes of β-1,3-glucanase and resistance of near-isogenic maize inbred lines to exserohilum turcicum [J]. Can. J. Plant Pathol, 1992, 14:285-290
[37] Reuveni R. Biochemical markers for disease resistance [M]. In: Molecular Methods in Plant Pathol., 1994, 99-114.
[38] Gu WK, Weeden NF, Yu J, et al. Large-scale cost effective screening of PCR products in marker-assisted selection application [J]. Theor. Appl. Genet., 1995, 91:465-470.
[39] Chunwongse J, Martin GB, Tanksley SD. Pre-germination genotypic screening using PCR amplification of half seeds [J]. Theor. Appl. Genet., 1993, 86:694-698.
[40] Knapp SJ. Marker assisted selection as a strategy for increasing the probability of selecting superior genotypes [J]. Crop Sci., 1998, 38:1164-1174.
[41] 陈升，张启发．分子标记辅助选择改良杂交水稻的白叶枯病抗性[J]．华中农业大学学报，2000，19：183-189．
[42] Mohan M, Nair S, Bhagnat A et al. Genome mapping, molecular markers and marker-assisted selection in crop plants [J]. Mol. Breeding, 1997, 3:87-103.
[43] Vos P, Hogers R, Bleekor M, et al. AFLP: a new technique for DNA fingerprinting[J]. Nucleic Acids Research, 1995, 23:4407-4414.
[44] Konieczny, A., and F.M. Ausubel. 1993. A procedure for mapping Arabidopsis mutations using co-dominant ecotype-specific PCR-based markers [J]. Plant J. 4:403-410.
[45] Akopyanz, N., N.O. Bukanov, T. Ulf Westblom, and D.E. Berg. 1992. PCR-based RFLP analysis of DNA sequence diversity in the gastric pathogen Helicobacter pylori [J]. Nucleic Acid Res. 20:6221-6225.
[46] Sachidanandam R .Weissman D ,Schmidt SC et al. A map of human genome sequence variation containing 1. 42 million single nucleotide polymorphisms [J ]. Nature, 2001, 409:928 - 933.
[47] McCouch SR, Kochert G, Yu ZH, et al. Molecular mapping of rice chromosome [J]. Theor Appl Genet 1988, 76:815-82.
[48] Harushima Y, Kurata N, Yano M, et al. Detection of segregation distortions in an indica-japonica rice cross using a high resolution molecular map [J]. Theor Appl Genet, 1996, 92:145-15.
[49] Giraudat J, Hauge BM, Valonc C. Isolation of the Arabidosis ABB gene by positional cloning [J]. Plant Cell. 1992,4:1251-1261.
[50] Arondel V, Lemieux B, Hwang I, et al. Map-based cloning of a gene controlling omega-3 fatty acid desaturation in Arabidosis [J]. Science, 1992, 258:1353-1355.
[51] Martin GB, Brommons H, Chumwongse J, et al. Map-based cloning of a protein kinase gene conferring disease in tomato [J]. Science, 1993, 262:1432-1436.
[52] Whitham S, Dinesh-Kumar SP, Choi D. The product of the tobacco mosaic virus resistance gene N: similarity to toll and the interleukin-1 receptor [J]. Cell, 1994, 78:1101-1115.
[53] Jones DA, Thomas CM, Hammond-Kosack KE. Isolation of the tomato C/-9 gene for resistance to Cladosporium fulvum by transposon tagging [J]. Science, 1994, 266:789-793.
[54] Lawrence GJ, Finnegen EJ, Ayliffe MA. The L6 gene for flax rust resistance is related to the Arabidopsis bacterial resistance gene RPS2 and the tobacco viral resistance gene N [J]. Plant Ceil, 1995,7:1195-1206.
[55] Liang P, Parde AB. Different display of eukaryotic messenger RNA by means of the polymerase chain reaction[J]. Science, 1992, 257(140):967-971.
[56] Diatchenlo L, lau YC, Campbell AP. Suppression subtractive hybridization: a method for generating differentially regulated or tissue - specific cDNA probes and libraries [J]. Proc Natl Acad Sci USA, 1996,93:6025-6030.
[57] Lisitsyn N, Wigler M. Cloning the differences between two complex genomes [J]. Science, 1993, 259:946-951.
[58] Johal GS, Briggs SP. Reductase activity encoded by the HM1 disease resistance gene in maize [J]. Science, 1992,258:985-987.
[59] Cai D, Kleine M, Kifle S, Haeioff HJ. Positional cloning of a gene for nematode resistance in sugar beet [J]. Science, 1997, 275:832-834.
[60] Hammond-Kosack KE, Jones JD. Plant disease resistance gene [J]. Ann Rev Plant Phyciol Plant Mol Biol, 1997,48:575-607.
[61] Dixon MS, Jones DA, Keddie JS. The tomoto Cf-2 disease resistance locus comprise two functional genes encoding leucine-rich repeat protein [J]. Cell, 1996, 84:451-459.
[62] Bent AF, Kankel BN, Dahlbeck D. RPS2 of Arabidopsis thaliana: a leuine-rich repeat class of plant disease resistance genes [J]. Science, 1994, 265:1856-1860.
[63] Grant MR, Godiard L, Sreaube E. Structure of the Arabidopsis RPM1 gene enable dual specificity disease resistance [J]. Science, 1995, 269:843-846.
[64] Laurent N, Tracey LM, Erik A, Martin P, et al. Pronounced intraspecific haplotype fivergence at the RPP5 complex disease resistance locus of arabidopsis [J] Plant Cell. 1999, 11:2099-2111.
[65] Catherine F, Gabriele S, Bent K. Molecular cloning of a new receptor-like kinase gene encoded the Lr10 disease resistance locus of wheat [J]. The Plant J, 1997,11(1): 45-52.
[66] Song WY, Wang GL, Chen LL. A receptor kinase-like protein encoded by the rice disease resistance geneXa21 [J]. Science, 1995,270:1804-1806.
[67] Kanazin V, Marek LF, Shoemker R. Resistance gene analogues are conserved and clustered in soybean [J]. Pro Natl Acad Sci USA, 1996, 93:11746-11750.
[68] Leister D, Ballvora A, Salamin F. A PCR-based approach for isolating pathogen resistance genes from potato with potential for wide application in plants [J]. Nature Genet, 1996, 13:421-429
[69] Mago, R., Nair, S., and Mohan, M. Resistance gene analogues from rice: cloning, sequencing and mapping [J]. Theor. Appl. Genet. 1999, 99: 50-57.
[70] Seah, S., Sivasithamparam, K., Karakousis, A., and Lagudah, E.S. Cloning and characterisation of a family of disease resistance gene analogs from wheat and barley [J]. Theor. Appl. Genet. 1998, 97: 937-945.
[71] Spiclmeyer, W., Robertson, M., Collins, N., Leister, D., SchulzeLefert, P., Seah, S., Moullet, O., and Lagudah, ES. A superfamily of disease resistance gene analogs are located on all homoeologous chromosome groups of wheat (Triticum aestivum) [J]. Genome, 1998,41: 782-788.
[72] Collins NC, Webb CA, Seah S. et al. The isolation and mapping of disease resistance gene analogs in maize [J]. Molecular Plant-Microbe Interactions, 1998, 11:968-978.
[73] Speulman E, Bouchez D, Holub EB, et al. Disease resistance gene homologs correlate with disease resistance loci of Arabidopsis thaliana[J]. The Plant J. 1998, 14(4):467-474.
[74]薛勇彪，唐定中，张燕生等．水稻基因组中R类抗病基因同源序列的分离[J]．科学通报，1998，43：277-281．
[75]王石平，刘克德，王江等．用同源序列的染色体定位寻找水稻抗病基因DNA片段[J]．植物学报，1998，40：42-50．
[76]Yu YG, Buss GR, Maroof MA. Isolation of a superfamily of candidate disease-resistance genes in soybean based on a conserved nucleotide-binding site [J]. Pro Natl Acad Sci USA. 1996, 93:11751-11756.
[77]Leister D, Kurth T, Schulze P. RFLP and physical mapping of resistance gene homologues in rice (O. sativa) and Barley (H. vulgare) [J]. Theor Appl genet, 1999, 98: 509-520.
[72]Creusot F, Macadre C, Cana EF, et al. Cloning and molecular characterization of three members of the NBS-LRR subfamily located in the vicinity of the Co-2 locus for anthracnose resistance in phaseolus vulgaris[J]. Genome, 1999, 42: 254-264.
[73]郑先武，翟文学，李晓兵等．水稻NBS-LRR类R基因同源序列[J]．中国科学(C辑)，2001，31(1)：43-50．
[74]李子银，陈受宜．水稻抗病基因同源序列的克隆、定位及表达[J]．科学通报，1999，44(7)：727-733．
[75]Katherine AS, Blake CK, Nurul MF, et al. Resistance gene candidates identified by PCR with degenerate oligonucleotide primers map to clusters of resistance gene in lettuce [J]. MIPI, 1998,11(8) 815-823.
[76]Mark GMA, Bas TLH, Eric BH, et al. Identification of R-gene homologous DNA fragments genetically linked to disease resistance loci in Arabidopsis thaliana [J]. MPMI, 1998, 11(4): 251-258.
[77]Lili Malekia, Justin DF, Robert L, Bowdenc, Bikram SG and John P. Physical and genetic mapping of wheat kinase analogs and nbs-lrr resistance gene analogs [J]. Crop Science, 2003, 43:660-670.
[78]Chen XM, Line RF, Leung H. Genome (?)anning for resistance-gene analogs in rice, barley and wheat by high resolution electrophoresis[J]. Theor Appl Genet. 1998, 97:345-355.
[79]Catherine F, Christophe R, Per K, et al. Molecular characterization of a new type of receptor-like kinase gene family in wheat. Plant Molecular Biology, 1998, 37:943-953.
[80]Ohmori T, Murata M, Motoyoshi F. Characterization of disease resistance gene-like sequences in near-isogenic lines of tomato. Theor Appl Genet. 1998, 96:331-338.
[81]Katherine AS, Blake CK, Nurul MF, et al. Resistance gene candidates identified by PCR with degenerate oligonuclcotide primers map to clusters of resistance gene in lettuce MIPI, 1998,11(8) 815-823.
[82]Pflieger S, Lefebvre V, Caranta C, et al. Disease resistance gene analogs as candidates for QTLs involved in pepper pathogen interactions[J] . Genome, 1999,42:1100-1110.
[83] Aarts MGM, Lintel HB, Holub EB, et al Identification of R-gene homologous DNA fragments genetically linked to disease resistance loci in Arabidopsis thaliana[J] Mol. Plant-Microbe Interact, 1998, 11(4): 251-258.
[84] Shen KA, Meyers BC, Islam-Faridi MN, et al. Resistance gene candidates identified by PCR with degenerate oligonucleotide primers map to clusters of resistance gene in lettuce [J]. Mol. Plant-Microbe Interact, 1998,11:815-823.
[85] Deng Z, Huang S, Ling P, et al. Cloning and characterization of NBS-LRR class resistance-gene candidate sequences in citrus [J]. Theor Appl Genet. 2000, 101:814-822.
[86] Leister D, Kurth J, Laurie DA, et al. Rapid reorganization of resistance gene homologues in cereal genomes [J]. Pro Natl Acad Sci USA. 1998, 95:370-375.
[1] Yu YG, Buss GR, and Saghai Maroof MA. Isolation of a superfamily of candidate disease-resistance genes in soybean based on a conserved nucleotide-binding site [J]. Proc Natl Acad Sci USA. 1996, 93(21):11751—11456.
[2] Blake C. Meyers, Alexander Kozik, Alyssa Griego, Hanhui Kuang, and Richard W. Michelmore. Genome-Wide Analysis of NBS-LRR-Encoding Genes in Arabidopsis [J]. Plant Cell, 2003, 15:809-834
[3] Spielmeyer W, Robertson M., Collins N, Leister D, Schulze LP, Seah S, Moullet O and Lagudah ES. A superfamily of disease resistance gene analogs are located on all homoeologous chromosome groups of wheat (Triticum aestivum) [J]. Genome. 1998, 41: 782-788.
[4] Collins, NC, Webb CA, Seah S, Ellis JG, et al. The isolation and mapping of disease resistance gene analogs in maize[J]. Molecular Plant-Microbe Interactions 1998, 11:968-978.
[5] Venancio S, Arahanaa, George L. Graefa, James E. Spechta, James R. Steadmanb and Kent M. Eskridge. PCR Markers for Triticum speltoides leaf rust resistance Gene Lr51 and their use to develop isogenic hard red spring wheat lines [J]. Crop Sci.2005, 45: 728-734.
[6] Lili Maleki, Justin D. Faris, Robert L. Bowden, Bikram S. Gill and John P. Fellers. Physical and Genetic Mapping of Wheat Kinase Analogs and NBS LRR Resistance Gene Analogs [J]. Crop Sci. 2003, 43:660-670.
[7] Seah S, Sivasithamparam K, Karakousis A, et al. Cloning and characterization of a family of disease resistance gene analogs from wheat and barley [J]. Theor Appl Genet. 1998, 97:937-945.
[8] Yuichi Tada. PCR-amplified resistance gene analogs link to resistance loci in rice [J]. Breeding Science. 1999, 49:267-273.
[9] Leister D, Ballvora A., Salamini F, and Gebhart C. A PCR-based approach for isolating pathogen resistance genes from potato with potential for wide application in plants [J]. Nat. Genet. 1996.14:421-429.
[10] Feuillet C, Travella S, Stein N, Albar L, Nublat A, Keller B. Map-based isolation of the leaf rust disease resistance gene Lr10 from the hexaploid wheat (Triticum aestivurn L.) genome [J]. Proc. Natl. Acad. Sci. USA 2003, 100:15253-15258.
[11] Aarts MGM, lintel Hekkert B, Holub EB, Beynon JK, Stiekema WJ, Pereira A. Identification of R-gene homologous DNA fragments genetically linked to disease resistance loci in Arabidopsis thaliana[J], Mol Plant Microbe Intertance. 1998, 11:251-258
[12] 薛勇彪，唐定中，张燕生等．水稻基因组中R类抗病基因同源序列的分离[J]．科学通报，1998，43：227-281．
[13] 王石平，刘克德，王江等．用同源序列染色体定位寻找水稻抗病基因DNA片段[J]．植物学报，1998，40(1)：42-50．
[14] 郑先武，翟文学，李晓兵等．水稻NBS—LRR类R基因同源序列[J]．中园科学(C辑)，2001，31(1)：43-50．
[15] 李子银，陈受宜．水稻抗病基因同源序列的克隆、定位及表达[J]．科学通报．1999，44(7)：727-733
[16] Shen KA, Meyers BC, Islam-Faridi MN, Chin DB, Stelly DM, and Michelmore RW. Resistance gene candidates identified by PCR with degenerate oligonucleotide primers map to clusters of resistance genes in lettuce[J]. Mol. Plant-Microbe Interact. 1998, 11, 815-823.
[17] Meyers BC, Dickerman AW, Michelmore RW, Sivaramakrishnan S, Sobral BW, and Young, ND Plant disease resistance genes encode members of an ancient and diverse protein family within the nucleotide-binding superfamily[J]. Plant J. 1999, 20, 317-332.
[18] Bai J, Pennill LA., Ning J, Lee SW, Ramalingam J, Webb CR, Zhao B, Sun Q, Nelson JC, Leach JE, and Hulbert SH. Diversity in nucleotide binding site-leucine-rich repeat genes in cereals[J]. Genome Res. 2002, 12:1871-1884.
[19] Richly, E., Kurth, J., and Leister, D. Mode of amplification and reorganization of resistance genes during recent Arabidopsis thaliana evolution [J]. Mol. Biol. Evol. 2002, 19: 76-84.
[20] Donald T., Pellerone, F., Adam-Blondon, AF, Bouquet A, Thomas M, and Dry I. Identification of resistance gene analogs linked to a powdery mildew resistance locus in grapevine [J]. Theor. Appl. Genet. 2002, 104: 610-618.
[21] Melotto M, Kelly JD. Fine mapping of the Co-4 locus of common bean reveals a resistance candidate, COK-4, that encodes for a protein kinase [J]. Theor. Appl. Genet. 2001,103:508-517.
22. Seah S, Bariana H, Jahier J, Sivasithamparam K, Lagudah. EST introgressed segment carrying rust resistance genes Yr17, Lr37 and Sr38 wheat can be assayed by a cloned disease resistance gene-like sequence[J]. Theor. Appl. Genet. 2001, 102:600-605.
[23] Madsen LH, Collins NC, Rakwalska M, Backes G, Sandal N, Krusell, Jensen J, Waterman EH, Jahoor A, Ayliffe M, Pryor A, Langridge P, Schulze-Lefert P, Stougaard J. Barley disease resistance gene analogs of the NBS-LRR class: identification and mapping[J]. Mol Genet Genomics 2003, 269:150-161.
[24] Meyers BC, Chin DB, Shen KA, Sivaramakrishanan S, Lavelle DO, Zhang ZM, Michelmore RW. The major resistance gene cluster in lettuce is highly duplicated and spans several megabases[J]. Plant Cell, 1998, 10:1817-1832.
[25] Huettel B, Santra D, Muelbauer FJ, Kahl G. Resistance gene analogues of chickpea (Cicer arietinum L.): isolation, genetic mapping and association with a Fusarium resistance gene cluster [J]. Theor. Appl. Genet. 2002, 105:479-490.
[26] Joseph Sambrook, David W. Russell. Preparation of plasmid DNA by alkaline lysis with SDS: Minipreparation. In: Molecular Cloning, third edition. 2001 by cold spring harbor laboratory press, cold spring harbor, New York..
[27] Newman T, Bruijn F J, Green P, Keegstra K, Kende H, McIntosh L, Ohirogge J, Raikhel N, Somerville S, Thomashow M, Retzel E, and Somerville CG. A summary of methods for accessing results from large-scale partial sequencing of anonymous arabidopsis cDNA clones[J], Plant Physiology, 1994, 106: 1241-1255.
[28] Baker B, Zambryski P, Staskawicz B, Dinesh-Kumar SP. Signaling in plant-microbe interactions. Science. 1997,276:726-733.
[29] Milligan SB, Bodeau J, Yaghoobi J, Kaloshian I, Zabel P and Williamson VM. The root knot nematode resistance gene Mi from tomato is a member of the leucine zipper, nucleotide binding, leucine-rich repeat family of plant genes. Plant Cell, 1998, 10:1307-1319.
[30] Rossi M, Goggin FL, Milligan SB, Kaloshian I, Ullman DE, Williamson VMThe nematode resistance gene Mi of tomato confers resistence against the potato aphid. Proc Natl Acad Sci USA 1998,95:9750-9754.
[31] Mago, R., Nair, S., and Mohan, M. Resistance gene analogues from rice: cloning, sequencing and mapping. Theor. Appl. Genet. 1999, 99: 50-57.
[32] Leister D, Kurth T, Schulze P. RFLP and physical mapping of resistance gene homologues in rice (O. sativa) and Bailey (H. vulgare)[J]. Theor Appl genet, 1999, 98:509-520.
[34] Diatchenlo L, lau YC, Campbell AP. Suppression subtractive hybridization: a method for generating differentially regulated or tissue - specific cDNA probes and libraries [J]. Proc Natl Acad Sci USA, 1996, 93:6025-6030.
[35] Catherine F, Gabriele S, Bent K. Molecular cloning of a new receptor-like kinase gene encoded the Lr10 disease resistance locus of wheat [J]. The Plant J, 1997,11(1): 45-52.
[36] Ohmori T, Murata M, Motoyoshi F. Characterization of disease resistance gene-like sequences in near-isogenic lines of tomato [J]. Theor Appl Genet. 1998,96:331-338.
[1] Hayward MD, Mcadam NJ, Jones JG, Evans C, Evans GM, Forster JM, Ustin A, Hossain KG, Quader B, Stammers M, Will JK. Genetic markers and the selection of quantitative traits in forage grasses [J]. Euphytica. 1994, 77:269-275.
[2] 余四斌，张启发．基因图谱在水稻遗传育种研究中的应用[M]．见：洪国藩主编．水稻基因组工程．上海：上海科学技术出版社．1999，79-116．
[3] 贾建航，李传友，邓启云，王斌．用AFLP标记快速构建遗传连锁图谱并定位一个新基因tms5[J]．Acta Botanica Sinica(植物学报：英文版)2003，45(5)：614-620．
[4] 吴茂清 张献龙 聂以春 贺道华．四倍体栽培棉种产量和纤维品质性状的QTL定位[J]．遗传学报．2003，30(5)：443-452．
[5] 黄烈健，杨俊品．玉米RFLP连锁图谱构建及大斑病QTL定位[J]．遗传学报．2002，29(12)：1100-1104．
[6] Hutchinson, J., Rees, H, and Seal, A. G. An assay of the activity of supplementary DNA in Lolium [J]. Heredity. 1979, 43:411-421.
[7] Warnke S. E., Barker R. E., Geunhwa Jung, Sung-Chur Sim. Genetic linkage mapping of an annual×perennial ryegrass population[J]. Theor Appl Genet, 2004, 109:294-304.
[8] Jones ES, Dupal MD. Dumsday JL, Hughes LJ, Forster JW. An SSR-based genetic linkage map for perennial ryegrass (Lolium perenne) [J]. Theor Appl Genet, 2002a,102:405-415.
[9] Bert PF, Charmer G, Sourdille P, Hayward MD, Balfourier F A high-density molecular map for ryegrass (Lolium perenne) using AFLP markers[J]. Theor Appl Genet, 1999, 99:445-452.
[10] Inoue M, Gao Z, Hirata M, Fujimori M, Cai H. Construction of a high-density linkage map of Italian ryegrass (Lolium multiflorum Lam.) using restriction fragment length polymorphism, amplified fragment length polymorphism, and telomeric repeat associated sequence markers [J]. Genome, 2004, 47:57-65.
[11] Brian P. Horgan and Fred H. Yelverton. Removal of perennial ryegrass from overseeded bermudagrass using cultural methods [J]. Crop Sci. 2001, 41: 118-126.
[12] Kevin B. Jensen, Kay H. Asay, and Blair L. Waldron. Dry matter production of orchardgrass and perennial ryegrass at five irrigation levels [J].Crop Sci. 2001, 41: 479-487.
[13] 杨中艺，陈三有．“黑麦草—水稻”草田轮作系统应用效益初探(案例研究)[J]．草业科学．1997，14(6)：35-39．
[14] 杨中艺，杨卓睿．稻田冬种黑麦草对后作水稻生长的影响及其机理初探[J]．草业科学．1997，14(4)：20-24．
[15] Golembiewski, R.C., T.K. Danneberger, and P.M. Sweeney. Potential of RAPD markers for use in the identification of creeping bentgrass cultivars [J]. Crop Sci. 1997, 37:212-214.
[16] Murry M, Thompson W F. Rapid isolation of high molecular weight plant DNA [J]. Nucl Acid Res, 1980, 8:668-673.
[17] Akkaya, M.A., Bhagwat A., and Cregan P.B. Length polymorphisms of simple sequence repeat DNA in soybean [J]. Genetics. 1992, 132:131-1139.
[18] Vos P, Hogers R, Bleekers M, Reijans M, van der Lee T, Hornes M, Frijters A, Pot J, Peleman J, Kuiper M, Zabesu M. AFLP: a new technique for DNA fingerprinting [J]. Nucleic Acids Res. 1995, 23:4407-4414.
[19] Myburg AA, Remington DL, O'Malley M, et al. High-throughput AFLP analysis using infraed dye-labeled primers and an automated DNA sequencer [J]. BioTechniques. 2001, 30:348-357.
[20] Haldane, J B S, The combination of linkage values and the calculation of distances between the loci of linked factors [J]. J. Genett. 1919, 8:299-309.
[21] Voorrips, R.E. MapChart: software for the graphical presentation of linkage maps and QTLs[J]. J. Hered. 2002,93:77-78.
[22] Forster JW, Jones ES, Ko" lliker R, Drayton MC, Dumsday J, Dupal MP, Guthridge KM, Mahoney NL, van Zijll de Jong E, Smith KF. Development and implementation of molecular markers for forage crop improvement. In: Spangenberg G (ed) Molecular breeding of forage crops [J]. Kluwer, Dordrecht, 2001, pp 101-133.
[23] Wu KS, Tanksley SD. Abundance polymorphism and genetic mapping of microsatellite in rice [J]. Mol Gen Genet. 1993, 241:225-235.
[24] Vos P, Hogers R, Bleeker M, Reijans M, Hornes M, Fritjers A, PotJ, Poleman J, Kuiper M and Zabeau M. AFLP: A new technique for DNA fingerprinting [J]. Nucreic Acid Res. 1995, 23: 4407-4414.
[25] Zhu J, Gale MD, Quarrie S, Jackson MT and Bryan GJ. AFLP markers for the study of rice biodiversity[J]. Theor. Appl. Genet. 1998, 96:602-611.
[26] Abdalla AM, Reddy OUK, El-zik KM and Pepper AE. Genetic diversity and relationships of diploid and tetraploid cottons revealed using AFLP[J]. Theor. Appl. Genet. 2001, 102: 222-229.
[27] Tohme J, Gonzalez DO, Beebe S and Duque MC. AFLP analysis of gene pools of a wild bean core collection [J]. Crop Sci. 1996, 36:1375-1384.
[28] Larson SR, Waldron BL, Monsen SB, John LS, Palazzo AJ, McCracken CL and Harrison RD. AFLP variation in agamospermous and dioecious bluegrasses of Western North America [J]. Crop Sci. 2001,41:1300-1305.
[29] Ferdinandez YSN, and Coulman BE. Evaluating genetic variation and relationships between two bromcgrass species and their hybrid using RAPD and AFLP markers [J]. Euphytica 2002, 125:281-291.
[30] Boukar O, Kong L, Singh BB, Murdock L and Ohm HW. AFLP and AFLP-derived SCAR markers associated with striga gesnerioides resistance in cowpea [J]. Crop Sci. 2004, 44: 1259-1264.
[31] Cooley MB, Pathirana S, Wu HJ, Kachroo P and Klessig DF. Members of the Arabidopsis HRT/RPP8 family of resistance genes confer resistance to both viral and omycete pathogens [J]. Plant Cell. 2000, 12,663-676.
[32] Aarts MG, Hekkert B, Holub EB, Beynon JL, Stiekema WJ and Pereira A. Identification of R gene homologous DNA fragments genetically linked to disease resistance loci in Arabidopsis thaliana [J]. Mol. Plant-Microbe Interact. 1998, 11, 251-258.
[33] Donald T, Pellerone F, Adam-Blondon A-F, Bouquet A, Thomas M and Dry I. Identification of resistance gene analogs linked to a powdery mildew resistance locus in grapevine [J]. Theor. Appl. Genet. 2002, 104,610-618.
[34] Meyers BC, Kozik A, Griego A, Kuang H and Michelmore RW. Genome-wide analysis of NBS-LRR-encoding genes in Arabidopsis [J]. The Plant Cell, 2003, 15:809-834.
[35] Kanazin V, Marek LF and Shoemaker RC. Resistance gene analogs are conserved and clustered in soybean [J]. Proc. Natl. Acad. Sci. USA. 1996, 93:11746-11750.
[1] Sumida Y, Fukubara H, Inoue T, Kajihara H, Mizuno K, Fujiwar T A study of the occurrence of dieback on Italian ryegrass (Lolium multiflorum) caused by Pyricularia sp.—Investigation of facts about occurrence and development of a new resistance test method on breeding [J]. Bull Yamaguchi Agric Exp Stn. 2003, 54: 29-36.
[2] Bain DC, Patel BN and Patel MV. Blast of ryegrass in Mississippi [J]. Plant Dis. Rep. 1972, 56:210.
[3] Carver, RB., Rush MC and Lindberg GD. An epiphytotic of ryegrass blast in Louisiana[J]. Plant Dis. Rep. 1972, 56:157-159.
[4] Viji G, Wu B, Kang S, Uddin W, Huff DR. Pyricularia grisea causing gray leaf spot of perennial ryegrass turf: population structure and host specificity[J]. Plant Dis. 2001, 85:817-826.
[5] Mizuno K, Yokohata Y, Oda T, Fujiwara T, Hayashi K, Ozaki R, Kobashi K, Ashizawa H, Ushimi T Breeding of a new variety 'Sachiaoba' in Italian ryegrass (Lolium multiflorum L.) and its characteristics[J]. Bull Yamaguchi Agric Expt Stn. 2003, 54:11-24.
[6] 辛国荣，岳朝阳．李雪梅等．“黑麦草-水稻”草田轮作系统的根际效应Ⅱ 黑麦草根系对土壤物理化学性状的影响[J]．中山大学学报(自然科学版)，1998，37(5)：78-82．
[7] 张新跃，何丕阳，何光武．“饲用玉米-黑麦草”草地农业系统的研究[J]．草业学报，2001，10 (2)：33-38．
[8] Bert PF, Charmet G, Sourdille P, Hayward MD, Balfourier F. A high-density molecular map for ryegrass (Lolium perenne) using AFLP markers [J]. Theor Appl Genet, 1999, 99: 445-452.
[9] Jones ES, Dupal MD, Dumsday JL, Hughes LJ, Forster JW. An SSR-based genetic linkage map for perennial ryegrass (Lolium perenne) [J]. Theor Appl Genet 2002a, 102:405-415.
[10] Hirata M, Fujimori M, Komatsu T. Development of simple sequence repeat (SSR) markers in Italian ryegrass [M]. In: 2nd international symposium on molecular breeding of forage crops, 19-24 November 2000, Lorne and Hamilton, Victoria, Australia.
[11] Inoue M, Gao Z, Hirata M, Fujimori M, Cai H. Construction of a high-density linkage map of Italian ryegrass (Lolium multifiorum Lam.) using restriction fragment length polymorphism, amplified fragment length polymorphism, and telomeric repeat associated sequence markers [J]. Genome, 2004, 47:57-65.
[12] Hayward MD, Forster JW, Jones JG, Dolstra O, Evans C, McAdam NJ, Hossain KG, Stammers M, Will J, Humphreys MO, Evans GM. Genetic analysis of Lolium. Ⅰ. Identification of linkage groups and the establishment of a genetic map [J]. Plant Breed, 1998, 117:451-455.
[13] Dumsday JL, Smith KF, Forster JW, Jones ES. SSR-based genetic linkage analysis of resistance to crown rust (Puccinia eoronata f. sp. lolii) in perennial ryegrass (Lolium perenne) [J]. Plant Pathol 2003, 52:628-637
[14] Curley J, Sim SC, Jung G, Leong S, Warnke S, Barker RE. QTL mapping of ryegrass blast resistance in ryegrass, and synteny-based comparison with rice blast resistance genes in rice [M]. In: Hopkins A, Wang ZY, Mian R, Sledge M, Barkers RE (eds) Molecular breeding of forage and turf. Kluwer, Dordrecht, 2004, pp 37-46.
[15] Fujimori M, Hayashi K, Hirata M, Mizuno K, Fujiwara T, Akiyama F, Mano Y, Komatsu T, Takamizo T. Linkage analysis of crown rust resistance gene in Italian ryegrass (Lolium multiflorum Lam.)[M]. In: Proceedings of plant and animal genomes Ⅺ conference, 11-15 January 2003, San Diego, p 46.
[16] Bert PF, Charmet G, Sourdille P, Hayward MD, Balfourier F. A high-density molecular map for ryegrass (Lolium perenne) using AFLP markers [J]. Theor Appl Genet. 1999, 99: 445-452.
[17] Michelmore R.W., Paran I., and Kesseli R.V. Identification of markers linked to disease-resistance genes by bulked segregant analysis: A rapid method to detect markers in specific genomic regions by using segregating populations [J]. Proc. Natl. Acad. Sci. USA 1991, 88:9828-9832.
[18] Blake C. Meyers, Alexander Kozik, Alyssa Griego, Hanhui Kuang, and Richard W. Michelmore. Genome-Wide Analysis of NBS-LRR-Encoding Genes in Arabidopsis[J]. Plant Cell, 2003, 15:809-834.
[19] Yu YG, Buss GR, Maroof MA. Isolation of a superfamily of candidate disease-resistance genes in soybean based on a conserved nucleotide-binding site [J]. Proc. Natl. Acad. Sci. USA, 1996, 93:11751-11756.
[20] Collins NC, Webb CA, Seah S, Ellis JG. et al. The isolation and mapping of disease resistance gene analogs in maize [J]. Molecular Plant-Microbe Interactions 1998, 11:968-978.
[21] Seah S, Sivasithamparam K, Karakousis A, et al. Cloning and characterization of a family of disease resistance gene analogs from wheat and barley [J]. Theor Appl Genet 1998, 97:937-945.
[22] Leister, D, Ballvora A, Salamini F, and Gebhart C. A PCR-based approach for isolating pathogen resistance genes from potato with potential for wide application in plants [J]. Nat. Genet. 1996, 14:421-429.
[23] Ikeda S, Takahashi W, Oishi H. Generation of expressed sequence tags from cDNA libraries of Italian ryegrass (Lolium multiflorum Lam.) [J]. Grassland Sci 2004, 49:593-598.
[24] Yuichi Tada. 1999. PCR-amplified resistance gene analogs link to resistance loci in rice [J]. Breeding Science 49:267-273.
[25] Murray MG, Thompson WF. Rapid isolation of high molecular weight plant DNA [J]. Nucleic Acids Research. 1980, 8: 4321-4325.
[26] 刘贤华，白晓军，胡川闽等．一种改良的质粒DNA的快述提取方法[J]．第三军医大学学报，1999，21(7)：531-532．
[27] Vos P, Hogers R, Bleekers M, Reijans M, van der Lee T, Hornes M, Frijters A, Pot J, Peleman J, Kuiper M, Zabesu M. AFLP: a new technique for DNA fingerprinting [J]. Nucleic Acids Res. 1995, 23:4407-4414.
[28] Myburg. AA, Remington DL, O'Malley DM, Sederoff RR, Whetten RW High-throughput AFLP analysis using, infrared dye-labeled primers and an automated DNA sequencer [J]. Bio-Techniques. 2001, 30:348-357.
[29] Van Ooijen JW, Voorrips RE. JoinMap 3.0 sof(?)vare for the calculation of genetic linkage maps [M]. Plant Research International, Wageningen, the Netherlands, 2001.
[30] Kosambi DD. The estimation of map distances from recombination values [J]. Ann Eugen, 1944, 12: 172-175.
[31] Konieczyny A, Ausubel FM. A procedure for mapping Arabidopsis mutations using co-dominant ecotype-specific PCR-based markers [J]. Plant J. 1993, 4:403-410.
[32] Drenkard E, Glazebrook J, Preuss D, Ausubel FM. Use of cleaved amplified polymorphic sequences (CAPS) for genetic mapping and typing [M]. In: Caetano-Anolles G, Gresshoff PM. DNA markers. Protocols, applications, and overviews [J]. Wiley-Liss, New York, 1997, pp 187-197.
[33] Williams MNV, Pande N, Nair S, Mohan M, Bennett J. Restriction fragment length polymorphism analysis of polymerase chain reation products amplified from mapped loci of rice (Oryza sativa L.) genomic DNA[J]. Theor Appl Genet. 1991, 82:489-498.
[34] Ghareyazie B, Huang N, Second G, Bennett J, Khush GS. Classification of rice3 germplasm. I. Analysis using ALP and PCR-based RFLP[J]. Theor Appl Genet. 1995, 91:218-227.
[35] Mano Y, Sayed-Tabatabaei BE, Graner A, Blake T, Takaiwa F, Oka S, Komatsuda T. Map construction of sequence-tagged sites (STSs) in barley (Hordeum Vulgare L.) [J]. Theor Appl Genet. 1999, 98:937-946.
[36] Talbert LE, Blake NK, Chee PW, Blake TK, Magyar GM. Evaluation of "sequence-tagged site" PCR products as molecular markers in wheat [J]. Theor Appl Genet. 1994, 87: 789-794.
[37] Meyers BC, Dickerman AW, Michelmore RW, et al. Plant disease resistance genes encode members of an ancient and diverse protein family within the nucleotide-binding super family [J]. Plant J., 1999, 20:317-332.
[38] Monosi B, Wisser RJ, Pennill L, Hullbert SH. Full-genome analysis of resistance gene homologues in rice [J]. Theor Appl Genet. 2004, 109:1434-1447.
[39] Zhou T, Wang Y, Chen JQ, Araki H, Jing Z, Jiang Z, Jiang K, Shen J, Tian D. Genome-wide identification of NBS genes in Japonica rice reveals significant expansion of divergent non-TIR NBS-LRR genes[J]. Mol Genet Genomics. 2004, 271:402-415.
[40] Lagudah ES, Moullet O, Appels R. Map-based cloning of a gene sequence encoding a nucleotide-binding domain and a leucine-rich region at the Cre3 nematode resistance locus of wheat [J]. Genome, 1997, 40:659-665.
[41] Yuichi Tada. PCR-amplified resistance gene analogs link to resistance loci in rice [J]. Breeding Science, 1999,49:267-273.
[42] Mago R, Nair S, Mohan M. Resistance gene analogues from rice: cloning, sequencing and mapping [J]. Theor Appl Genet. 1999,99: 50-57.
[43] Leister D, allvora A, Salamini F, et al. A PCR-based approach for isolating pathogen resistance genes from potato with potential for wide application in plants [J]. Nat. Genetics, 1996, 14:421-429.
[44] Yu YG, Buss GR, Maroof MA. Isolation of a superfamily of candidate disease-resistance genes in soybean based on a conserved nucleotide-binding site [J]. Proc. Natl. Acad. Sci. USA. 1996, 93:11751-11756.
[45] Wang Z, Taramino G, Yang D, et al. Rice ESTs with disease-resistance gene-or defence-response gene-like sequence mapped to regions containing major resistance genes or QTLs [J]. Mol Genet Genomics, 2001, 265:302-310.
[46] Moller MG, Taylor C, Rasmussen SK, Holm PB. Molecular cloning and characterization of two genes encoding asparagines synthetase in barley (Hordeum vulgare L.) [J]. Biochim Biophys Acta 2003, 1628:123-132
[47] Dumsday JL, Smith KF, Forster JW, Jones ES. SSR-based genetic linkage analysis of resistance to crown rust (Puccinia coronata f. sp. lolii) in perennial ryegrass (Lolium perenne) [J]. Plant Pathol 2003,52:628-637
[48] Mizuno K, Yokohata Y, Oda T, Fujiwara T, Hayashi K, Ozaki R, Kobashi K, Ashizawa H, Ushimi T. Breeding of a new variety 'Sachiaoba' in Italian ryegrass (Lolium multiflorum L.) and its characteristics (in Japanese) [J]. Bull Yamaguchi Agric Expt Stn 2003, 54:11-24.
[49] Tabien RE, Li Z, Paterson AH, Marchetti MA, Stansel JW, Pinson SRM. Mapping QTLs for field resistance to the rice blast pathogen and evaluating their individual and combined utility in improved varieties[J]. Theor Appl Genet. 2002, 105:313-324.
[50] Hayward MD, Mcadam NJ, Jones JG, Evans C, Evans GM, Forster JM, Ustin A, Hossain KG, Quader B, Stammers M, Will JK. Genetic markers and the selection of quantitative traits in forage grasses[J]. Euphytica 1994, 77:269-275.
[51] Curley J, Sim SC, Jung G, Leong S, Warnke S, Barker RE. QTL mapping of ryegrass blast resistance in ryegrass, and synteny-based comparison with rice blast resistance genes in rice[M]. In: Hopkins A, Wang ZY, Mian R, Sledge M, Barkers RE(eds) Molecular breeding of forage and turf. Kluwer, Dordrecht, 2004, pp 37-46.