摘要
楸树（Catalpa bungei C.A. Meyer）是优良的用材树种和园林观赏树种，广泛应用于建筑、家具及园林绿化等行业，其栽培应用受到越来越多的重视。但楸树自交不育，加之营养繁殖困难，导致其资源匮乏，阻碍了楸树的大面积发展。因此，研究楸树杂交育种，实现有性繁殖，有效的解决楸树发展受限等问题，为楸树的大面积推广利用奠定基础。本论文以楸树（包括连云港云台山楸树和南京老山林场楸树，简称CB-1和CB-2）和滇楸(Catalpafargesii Bur. f. duclouxii (Dode) Gilmour)（简称CF）为材料，对其花粉贮藏条件、影响其花粉萌发和花粉管生长的因素进行了比较研究，并对花粉壁及相关花器官的蛋白特性进行了电泳分析。利用荧光显微技术研究了楸树杂交授粉后花粉在柱头的萌发及花粉管在花柱中的生长动态，确定楸树的不亲和类型及特点，通过蛋白纯化技术对杂交后的花柱、子房蛋白进行了分离、纯化、鉴定，初步确定了与楸树自交不亲和相关的蛋白特性。本研究的主要结论如下：
1．花粉在25℃暗光条件下，培养6h后萌发效果最佳，此时萌发率超过80%，花粉管长度大于300μm。不同楸树花粉对低温及超低温的反应不同，而且随贮藏时间的延长，不同温度条件对花粉生活力的影响也不同，总体上表现为贮藏温度越低，花粉生活力变化就越小。-70℃和液氮条件适宜楸树花粉长期贮藏，-20℃或4℃条件适合短期贮藏，而常温条件下花粉生活力很容易丧失，不适合花粉的贮藏。在液氮条件下贮藏花粉时，最适宜的解冻方法是在35～38℃水浴条件下化冻5min。
2．离体条件下花粉萌发和花粉管生长受多种因素的影响。适宜的培养温度为24～28℃，当温度过高或过低时均会抑制花粉萌发及花粉管生长。蔗糖浓度、聚乙二醇（PEG-4000）浓度、pH值都在一定范围内对花粉萌发及花粉管生长起促进作用，而超过一定值后起抑制作用。适宜的培养基组分为：蔗糖浓度为15～25g·L~(-1)，聚乙二醇（PEG-4000）浓度为20g·L~(-1)，pH值为5.0～5.6。
3．花粉壁蛋白的提取宜选用超声波提取法，超声参数最适为：功率400W、超声时间3秒／次、间歇时间6秒／次、分三个回合共120次、每回合之间相隔5分钟。适宜的花粉蛋白提取液采用pH7.8的Tris-HCl。SDS-PAGE电泳分析表明，各树种共有蛋白包括：86.8kD、74.2kD、70.0kD、45.0kD、43.0kD、37.1kD、35.4kD、34.2kD、33.0kD、32.3kD、30.7kD、29.7kD、28.2kD、20.7kD、19.8kD、17.0kD、15.2kD、12.4kD等。特异蛋白包括CB-1和CB-2的53.8kD、23.4kD蛋白，CB-2和CF的38.5kD蛋白，CB-1和CF的26.5kD蛋白，CB-1独有的35.0kD蛋白。与SDS-PAGE对应的IEF-SDS-PAGE中的特异点包括：CB-1蛋白：分子量35kD，pI4.75；CF和CB-1蛋白：分子量为26.5kD，pI5.55。
4．花器官中的可溶性蛋白质含量表现为子房＞花萼＞花柱＞花瓣。SDS-PAGE电泳分析表明，花柱共有蛋白包括：96kD、45kD、32kD、29kD、28kD、27kD、17kD、16kD、13kD、12kD等；特异蛋白包括：CB-1的42kD蛋白，CB-2和CF的58kD、24kD蛋白，CB-2的64kD、19kD蛋白，CF的37kD蛋白。子房中共有蛋白包括：45kD、32kD、29kD、28kD、27kD、25kD、23kD、21kD、20kD、19kD、17kD、15kD、12kD等；特异蛋白包括CB-1的41kD、38kD、23kD蛋白。花萼中共有蛋白包括：72kD、45kD、38kD、37kD、32kD、30kD、29kD、28kD、24kD、23kD、22kD、21kD、20kD、18kD、17kD、15kD、14kD、13kD、12kD等；特异蛋白包括CB-1的88kD、65kD蛋白，CB-2和CF中的52kD、40kD、26kD蛋白。花瓣内蛋白条带较少，蛋白带主要集中在12～45kD之间，特异蛋白包括CB-1的63kD蛋白。
5．自、异交授粉后花粉在柱头上萌发的时间存在差异，自交萌发时间长于异交，自交需4～8h，异交2～3h。花粉管在花柱内的生长表现为自交花粉管生长速度缓慢，而异交花粉管生长迅速，异交授粉后大约40h即有花粉管伸长进入子房，而自交授粉后79～143h有花粉管进入子房，且部分花粉管在花柱1/3处生长受阻，花粉管在生长过程中会出现倒长、弯曲等异常现象，表现为明显的自交不亲和而异交亲和的特性。
6．自、异交花柱、子房的蛋白纯化分离后，自交花柱中获得5个主峰，异交花柱中获得4个主峰；而自交子房有2个主峰，异交子房有3个主峰。对各峰的蛋白质鉴定结果表明：自交花柱的S1（第一峰）、S2（第二峰）、S3（第三峰）峰的收集液中的蛋白明显抑制自花花粉萌发，主要蛋白组分包括S1峰的52kD、38kD蛋白，S2峰的52kD、44kD、38kD、29kD蛋白，S3峰的38kD、29kD蛋白；同时发现自交子房的S2（第二峰）峰蛋白也较强的抑制自花花粉萌发，蛋白组分为52kD、44kD、38kD、29kD蛋白。反映楸树自交后花柱、子房中均有抑制自花花粉管生长的蛋白，具有子房内晚期不亲和特点。
Catalpa bungei C.A. Meyer is a precious timber and ornamental species. The cultivationand application of Catalpa bungei have been paid attention more and more recently due to itswidely uses in architecture, furniture and landscape. However, the seedlings shortage of Catalpabungei resulting from its self incompatibility and difficulty in vegetative reproduction severelystunted the extensive development of Catalpa bungei plantation. Therefore, studying thecrossbreeding and sexual propagation of Catalpa bungei would be very helpful in supplying theseedling resources and in developing the Catalpa bungei plantations. The Catalpa bungei C.A.Meyer from Yuantai mountain of Lianyungang (CB-1), Catalpa bungei C.A. Meyer fromLaoshan Forest Farm of Nanjing (CB-2) and Catalpa fargesii Bur. f. duelouxii (Dode) Gilrnourfrom Nanjing Forestry University (CF) were selected to study the storage conditions of theirpollen and the factors affecting their pollen germination and pollen tube development in thisthesis. The electrophoretic analysis of specific proteins in the pollen wall and other floral organsof catalpa were also done. By the techniques of fluorescence microscopy, the dynamics of pollengermination in the stigma and pollen tube development in the style after cross-pollination werestudied and the self-incompatibility types and their traits of Catalpa bungei species wereidentified. The proteins in the style and in the ovary of Catalpa bungei after cross pollinationwere separated, purified and identified and the properties of the protein associated withself-incompatibility of Catalpa bungei were primarily explicated. The main results were showedas follow:
1. The optimal conditions for pollen germination of Catalpa bungei were culture in the darkand25℃for6h, at which the pollen germination rate was over80%and the pollen tube lengthwas over300μm. The responses of the pollen to low and ultralow temperature varied with theCatalpa bungei species and the influences of temperature conditions on pollen viability variedwith the storage time. It was found that when needed to be stored for a long term, the Catalpabungei pollen was feasibly stored in the liquid nitrogen frozen at the temperature of-70℃, whenneeded to be stored for a short term, the optimal storage temperature was-20℃or4℃. Thepollen could not be sorted at the room temperature at which its viability was easily lost. Theoptimal method of thawing for the frozen pollen in the liquid nitrogen was heating the pollen for5minutes at35-38℃in the electric-heated thermostatic water bath.
2. There were several factors affecting the Catalpa bungei pollen germination and the pollentube development under in vitro culture. The optimal temperature for the pollen in vitro culturewas24-28℃, both higher and lower than the optimal temperature could inhibit the pollengermination and the growth of pollen tube. The pollen germination and the growth of pollen tube were promoted when the concentration of sucrose and polyethylene glycol (PEG-4000) and pHin the medium were the lower while they were inhibited when the concentration of sucrose wasover15-25g·L~(-1), the concentration of PEG-4000over20g·L~(-1), and the pH value over5.0～5.6.
3. The ultrasonic extraction was the suitable for the protein in the pollen wall and theoptimal parameters were ultrasonic power of400W for3seconds at an interval of6seconds, andthe ultrasonication of120times were finished in3rounds at an interval of5minutes. Thesolution of Tris-HCl with a7.8pH was suitable for the extraction of the pollen protein. By thetechnique of SDS-PAGE, it was found that the common protein86.8kD,74.2kD,70.0kD,45.0kD,43.0kD,37.1kD,35.4kD,34.2kD,33.0kD,32.3kD,30.7kD,29.7kD,28.2kD,20.7kD,19.8kD,17.0kD,15.2kD, and12.4kD were existed in all three Catalpa bungei tree speciestested. However, the specific proteins of CB-1and CB-2were53.8kD and23.4kD, the specificprotein of CB-2and CF was38.5kD, and the specific protein of CB-1and CF was26.5kD. Thespecific protein35.0kD was found only in the CB-1species. In comparison with the analysis ofSDS-PAGE, the isoelectric point (pI) of the specific protein35.0kD in the CB-1species was4.75in the analysis of IEF-SDS-PAGE, and the pI of the specific protein26.5kD in CF and CB-1species was5.55.
4. The soluble protein content in the floral organs was in the order of ovary> calyx> style>petal. The analysis of SDS-PAGE indicated that the common proteins included96kD,45kD,32kD,29kD,28kD,27kD,17kD,16kD,13kD, and12kD in the style. The specific protein42kDwas found in the style of CB-1, the protein58kD and24kD was found in the CB-2and CF species,the64kD and19kD in the CB-2, and the37kD in the style of CF species. In the ovary, thecommon protein include protein45kD,32kD,29kD,28kD,27kD,25kD,23kD,21kD,20kD,19kD,17kD,15kD, and12kD, while the specific protein41kD,38kD, and23kD were found inCB-1species. In the calyx, the common proteins included protein72kD,45kD,38kD,37kD,32kD,30kD,29kD,28kD,24kD,23kD,22kD,21kD,20kD,18kD,17kD,15kD,14kD,13kD,and12kD, while the specific protein88kD and65kD were found in the CB-1, and the protein52kD,40kD, and26kD in CB-2and CF species. The protein bands was relative few in the petaland mainly distributed in12-45kD, and the specific protein63kD was found in the CB-1species.
5. The time required for the pollen germination in stigma was4-8h after self-pollinationwhile it was2-3h after cross-pollination. The growth of pollen tubes in cross-pollination wasfaster and only in40h after cross-pollination the pollen tubes were entered into the ovary.However, the growth of pollen tubes in self-pollination in style was slower and some tubes wereinterrupted in the1/3of style. In the79-143h after self-pollination there were some pollen tubesentered into the ovary, and the abnormal morphology of the pollen tubes such as bent, curved,and growing downward were observed during the pollen tubes growth. This indicated thatCatalpa bungei was of the characteristics of self-incompatibility and cross-compatibility.
6. The proteins in the style and in the ovary were separated and purified after self-pollinationand cross-pollination. Five main peaks were obtained from the style of self-pollination and four main peaks from the style of cross-pollination. While from the ovary, two main peaks wereobtained in self-pollination and three peaks in cross-pollination. The identification of proteinsfrom the peaks showed that the proteins form the peak S1(first peak), S2(second peak), and S3(third peak) in the styles of self-pollination had obvious inhibitions on the germination of selfpollen. The main protein components were52kD and38kD in the peak S1, while52kD,44kD,38kD and29kD were mainly existed in the S2and38kD and29kD in the S3. Meanwhile, theprotein52kD,44,38, and29were also identified in the ovary of self-pollination, and theseproteins had relatively strong inhibitions on the germination of self pollen either. These suggestedthat catalpa species was of the characteristic of late-acting ovarian self-incompatibility due to theproteins inhibited pollen tubes in the style and in the ovary after self-pollination.
引文
[1] Allen A M, Hiscock S J. Evolution and phylogeny of self-incompatibility systems in angiosperms, In:Self-incompatibility in flowering plants: evolution, diversity, and mechanisms[M]. Springer-VerlagBerlin,2008,73-101.
[2] Anderson M A, Cornish E C, Maus L, et al. Cloning of cDNA for a stylar glycoprotein associated withexpression of self-incompatibility in Nicotiana Alata[J]. Nature,1986,321:38-44.
[3] Antonia P, Solveig D L, Madina F, et al. Five gametophytic mutations affecting pollen development andpollen tube growth in Arabidopsis thaliana[J]. Genetics,2001,158:1773-1783.
[4] Bai C, Sen P, Hofmann K, et al. SKP1connects cell cycle regulators to the ubiquitin proteolysismachinery through a novel motif, the F-box[J]. Cell,1996,86(2):263-274.
[5] Barber J T, Steward F C. The proteins of Tulipa and their relation to morphogenesis[J]. Develop Biol,1968,17:326-349.
[6] Bateman A J. Self-incompability systems in angiosperms[J]. Heredity,1952,6:285-310.
[7] Beny A, Mary P, Mason P, et al. The effect of high temperature and high atmospheric CO2oncarbohydrate changes in bell pepper (Capsicum annuum) pollen in relation to its germination[J].Physiologia Plantarum,2001,112:505-512.
[8] Bower M S, Matias D D, Fernandes-Carvalho E, et al. Two members of the thioredoxin-h family interactwith the kinase domain of a Brassica S locus roceptor kinase[J]. Plant Cell,1996,8:1641-1650.
[9] Bradford M M. A rapid and sensitive method for the quantities of protein utilizing the principle of proteindye binding[J]. Anal Biochem,1976,72:248-252.
[10] Brewbaker J L. Biology of the angiosperm pollen grain[J]. Indian Journal of Genetics Plant Breeding,1959,19:121-133.
[11] Brewbaker J L, Kwack B H. The essential role of calciumion in pollen germination and pollen tubegrowth[J]. Amer J Bot,1963,50:859-865.
[12] Broothaerts W J, Andre vau Laere, Raf wiffers, et al. Purification and N-terminal sequencing of styleglycoproteins associated with self-incompatibility in Petunia hybria[J]. Plant Mol Biol,1989,14:93-102.
[13] Cabrillac D, Cock J M, Dumas C, et al. The S-locus receptor kinase is inhibited by thioredoxins andactivated by pollen coat proteins[J]. Nature,2001.410:220-223.
[14] Casselman A L, Vrebalo J, Conner J A, et al. Determining the physical limits of the brassica S locus byrecombinatioal analysis[J]. Plant Cell,2000,12:23-33.
[15] Certal A C, Sanchez A M, Kokko H, et al. S-RNases in apple are expressed in the pistil along the pollentub growth path[J]. Sex Plant Reprod,1999,12:94-98.
[16] Chen C H, Nasrallsh J B. A new crass of S sequences defined by a pollen recessive self-incompatibilityallele of Brassica oleracea[J]. Mol Gen Genet,1990,222:241-248.
[17] Clark K R, Okuley J J, Collins P D, et al. Sequence variability and developmental expression of S-allelesin self-incompatible and pseudo-self-compatible petunia[J]. Plant Cell,1990,2:815-826.
[18] Comish E C, Pettitt J M, Bonig L, et al. Developmentally controlled expression of a gene associated withself-incompatibility in Nicotiana alata[J]. Nature,1987,326:99-102.
[19] Cruz-Garcla F, Hancock C N, Kim D, et al. Stylar glycoproteins bind to S-RNase in vitro[J]. Plant J,2005,42:295-304.
[20] Dai S, Li L, Chen T, et al. Proteomic analyses of Oryza sativa mature pollen reveal novel proteinsassociated with pollen germination and tube growth[J]. Proteomics,2006,6(8):2504-2529.
[21] De Nettancourt D. Incompatibility in angiosperms[M]. Spinger-Verlag, New York.1977.
[22] De Nettancourt D. Incompatibility and incongruity in wild and cultivated plants[M]. Berlin: Springer-Verlag,2001.
[23] Dodds P N, Bonig I, Du H, et al. S-RNase gene of Nicotiana alata is expressed in developing pollen[J].Plant Cel1.1993,5(12):1771-1782.
[24] Dodds P N, Ferguson C, Clarke A E, et al. Pollen-expressed S-RNases are not involved in self-incompa-tibility in Lycopersicon peruvianum[J]. Sex Plant Reprod,1999,12(2):76-87.
[25] Doughty J, Dixon S, Hiscock S J, et al. PCP-A1defensin-like brassica pollen coat protein that binds the Slocus glyeoprotein, is the product of gametophytic gene expression[J]. Plant Cell,1998,10(8):1333-1347.
[26] East E M. The distrbution of self-sterility in the flowering plants[J]. Proc Am Philo Soc,1940,82(4):449-518.
[27] Edlund A F, Swanson R, Preuss D. Pollen and stigma structure and function: the role of diversity inpollination[J]. Plant Cell,2004,16(l): S84-S97.
[28] Entani T, Lwano M, Shiba H, et al. Comparative analysis of the self-incompatibility (S-) locus region ofPrunus mume: identification of a pollen-expressed F-box gene with allelic diversity[J]. Genes Cells,2003,8(3):203-213.
[29] Evans P T, Holaway B L, Malmberg R L. Biochemical differentiation in the tobacco flower probed withmonoclonal antibocies[J]. Planta,1988,175:259-269.
[30] Foote H G, Franklin-Tong V E, Walker E A, et al. Cloning and expression of a novel self-incompatibility(S-)gene from Papaver rhodes L[J]. Proc Natl Acad Sci USA,1994,91(6):2265-2269.
[31] Franklin-Tong V E, Lawrence M J, Franklin F C H. Characterization of a stigmatic component of fromPapaver rhoeas L. which exhibits the specific activity of a self-incompatibility (S-) gene product[J].New Phytol,1989,112(2):307-315.
[32] Franking T V E, Atwal K K, Howell E C, et al. Self-incompatibility in Papaver rhoeas: there is noevidence for the involvement of stigmatic ribonuclease activity[J]. Plant Cell and Environment,1991,14(4):423-429.
[33] Franklin W M. Staining and observing pollen tubes in the style by means of fluorescence[J]. Stain Tech,1959,34(3):125-128.
[34] Gasser C S. Molecular studies of the differentiation of floral organs[J]. Annu Rev Plant Mol Biol,1991,42:621-649.
[35] Gianazza. Isoelectric focusing[C]. Pharmacia Fine Chemical,1982,23-35.
[36] Gibbs P E, Bianchi M B. Does late-acting self-incompatibility (LSI) show family clustering? Two morespecies of Bignoniaceae with LSI: Dolichandra cynanchoides and Tabebuia nodosa[J]. Ann Bot,1999,84(4),449-457.
[37] Goldralj A, Kondo K, Lee C B, et al. Compartmentalization of S-RNase and HT-B degradation in self-incompatible Nicotiana[J]. Nature,2006,439:805-810.
[38] Golz J F, Su V, Clarke A E, et al. A molecular description of mutations afecting the pollen component ofthe Nicotiana alata S Locus[J]. Genetics,1999,152(3):1123-1135.
[39] Goring D R, Rothateig S J. The S-locus receptor kinasein in a self-incompatible Brassica napus lineencodes a functional serine/threonine kinase[J]. Plant Cell,1992,4(10):1273-1281.
[40] Goring D R, Glavin T L, Schafer U, et al. An S roceptor kinase gene in self-compatible Brassica napushas a1-bp deletion[J]. Plant Cell,1993,5(5):531-539.
[41] Goring D R, Walder J C. Self-injection-a new kinase connection[J]. Science,2004,303:1474-1475.
[42] Gray J E, McClure B A, Bonig I, et al. Action of the style product of the self-incompatibility gene ofNicotiana alata (S-RNase) on in vitro-grown pollen tubes[J]. Plant Cell,1991,3(3):271-283.
[43] Gu T, Mazzurco M, Sulaman W, et al. Binding of an arm repeat protein to the kinase domain of theS-locus receptor kinase[J]. Proc Natl Acad Sci USA, l998,95(1):382-387.
[44] Hanks S K, Quinn A M. Protein kinase catalytic domain sequence database: identification of conservedfeatures of primary structure and classification of family members[J]. Methods Enzymology,1991,200:38-62.
[45] Heslop-Harrison J, Heslop-Harrison Y. Evaluation of pollen viability by enzymatically inducedfluorescence; In tracellular hydrolysis of fluorescein diacetate[J]. Stain Tech,1970,45(3):115-120.
[46] Hesldp-Harrison J. Incompatibility and the pollen-stigma interation[J]. Ann Rev Plant Physiol,1975,26:403-425.
[47] Heslop-Harrison J. Pollen-stigma irteraction and cross-incompatibility in the grasses[J]. Science,1982,215(4538):1358-1364.
[48] Hiratuka S, Zhang S-L, Nakagawa E. et al. Selective inhibition of the growth of incompatible pollen tubesby S-protein in the Japanese pear[J]. Sex Plant Reprod,2001,13:209-215.
[49] Hiratsuka S, Tezuka T, Yamamoto Y. Analysis of self-incompatibility reaction in Easter lily by using heattreatments[J]. J Amer Soc Hort Sci,1989,114:505-508.
[50] Hiratsuka S, Kitoh Y, Matsurnshirna J. Induction of deformed pollen tube tips and their morphologicalcharacteristics in self-incompatible Japanese pear[J]. J Japan Soc Hort Sci,1991,60(2):257-265.
[51] Hiscock S J, Kues U, Diekinson H G. Molocular mechanisms of self-incompatibility in flowering plantsand fungi-diferent means to the same end[J]. Trends Cell Biol,1996,6:421-428.
[52] Holmes-Davis R, Tanaka C K, Venselw H, et al. Proteome mapping of mature pollen of Arabidopsisthaliana[J]. Protemomics,2005,5(18):4864-4884.
[53] Hua Z, Kao T H. Identification and characterization of components of a putative petunia S-LocusF-box-containing E3ligase complex involved in S-RNase-based self-incompatibility[J]. Plant Cell,2006,18(10):2531-2553.
[54] Huang S, Lee H S, Karunanandaa B, et al. Ribonuclease activity of Petunia inflata S proteins is essentialfor rejection of self-pollen[J]. Plant Cell,1994,6(7),1021-1028.
[55] Huang J, Zhao L, Yang Q, et al. AhSSK1, a novel SKP1-like protein that interacts with the S-locus F-boxprotein SLF[J]. Plant J,2006,46(5):780-793.
[56] Hulskamp M, Kopczak S D, Horejsi T F, et al. Identification of genes required for pollen-stigmarecognition in Arabidops thaliana[J]. Plant J,1995,8(5):703-714.
[57] Ikeda S, Nasrallah J B, Dixit R, et al. An aquaporin-like gene required for the Brassicaself-incomoatibility response[J]. Science,1997,276(5318):1564-1566.
[58] Ioerger T R, Gohlke J R, Xu B, et al. Primary structural features of the self-incompatibility protein insolanaceae[J]. Sex Plant Reprod,1991,4:81-87.
[59] Ishimizu T, Sato Y, Salto T, et al. Identification and partia amino acid sequences of seven S-RNasesassociated with self-incompatibility of the Japanese pear[J]. Pyrus pyriforia Nakai. J Biochem,1996,120(2):326-334.
[60] Ishimizu T, Norioka S, Nakanish T, et al. S-Genotype of Japanese Pear ‘Hosui’[J]. J Japan Soc Hort Sci,1998,67(1):35-38.
[61] Iwano M, Shiba H, Funato M, et al. Immunohistochemical studies on translocation of pollen S-haplotypedeterminant in self-incompatibility of Brassica rapa[J]. Plant Cell Physoil,2003,44(4):428-436.
[62] Jahnen W, Batterham M P, Clarke A E, et al. Identification, isolation, and N-terminal sequening of styleglycoproteins associated with self-incompatibity in Nicotiana alata[J]. Plant Cell,1989,1(5):493-499.
[63] Kachroo A, Schopfer C R, Nasrallh M E, et al. Allele-specific receptor-ligand interactions in Brassicaself-incompatibility[J]. Science,2001,293(5536):1824-1826.
[64] Kachroo A, Nasrallah M E, Nasrallah J B. Self-incompatibility in the Brassicaceae: Roceptor-ligandsignaling and cell-to-cell communication[J]. Plant Cell,2002,14: S227-S238.
[65] Kakeda K, Jordan N D, Conner A, et al. Identification of residues in a hydrophilic loop of the Papaverrhoeas S protein that play a crucial role in the recognition of incompatible pollen[J]. Plant Cell,1998,10(10):1723-1731.
[66] Kao T H, Mc Cubbin A G. How flowering plants discriminate between self and non-self pollen to preventinbreeding[J]. Proc Natl Acad Scj USA,1996,93(22):12059-12065.
[67] Kao T H, Tsukamoto T. The molecular and genetic bases of S-RNase-based self-incompatibi1iy[J]. PlantCell,2004,16(1): S72-S83.
[68] Kenrick J, Kaul V, Williams E G. Self-incompatibility in Acacia retinodes: Site of pollen-tube arrest is thenucellus[J]. Planta,1986,169,245-250.
[69] Kheyr-Pour A, Bintrim S B, loerger T R, et al. Sequence diversity of postil S-proteins associated withgametophytic self-incompatibility in Nicoriana alata[J]. Sex Plant Reprod,1990,3:88-97.
[70] Kipreos E T, Pagano M. The F-box protein family[J]. Genome Biol,2000,1(5):3001-3002.
[71] Knight R, Rogers H H. Incompatibility in Theobroma cacao[J]. Heredity,1955,9:69-77.
[72] Kondo K, Yamamoto M, Itahashi R. Insights into the evolution of self-compatibility in Lycopersicon froma study of stylar factors[J]. Plant J,2002,30(2):143-153.
[73] Kotlunowa M, Turetimer J, Cox K H, et al. Different temporal and spatial gene expression patterns occurduring another development[J]. Plant Cell,1990,2(12):1201-1224.
[74] Kowyama Y, Kunz C, Lewis I, et al. Self-compatibility in a lycopersicon peruvianum variant (LA2157) isassociated with a lack of style S-RNase activity[J]. Theor Appl Gen,1994,88:859-864.
[75] Kurup S, Ride J P, Jordan N, et al. Identification and cloning of related self-incompatibility S-genes inPapaver rhoeas and Papaver nudiceule[J]. Sex Plant Reprod,1998,11(4):192-198.
[76] Lai Z, Ma W, Han B, et al. An F-box gene linked to the self-incompatibility (S)locus of Antirrhium isexpressed specifically in pollen and tapetum[J]. Plant Mol Biol,2002,50(1):29-42.
[77] Lange J H. Pollen tube growth in citrus[J]. Agroplatae,1973,5(3):73-77.
[78] Lee H S, Huang S, Kao T H. S proteins control rejection of incompatible pollen in Petunia inflata[J].Nature,1994,367:560-563.
[79] Lewis D. Serological reactions of pollen incompatibility substances[J]. Proc Roy Soc Lond B,1952,140(898):127-135.
[80] Lind J L, Bonig I, Clarke A E, et al. A style-specific120-kDa glycoprotein enters pollen tubes ofNicotiana alata in vivo[J]. Sex Plant Reprod,1996,9:75-86.
[81] Lizuka M. Studies on the fertility of artificial polyploidy plants Ⅳ Dual pollination with self-incompatibleand cross-compatible pollen in Brassica[J]. Bul Res Inst Food, Sci Kyoto U-niv,1957,19:52-62.
[82] Ludmila R, Eva H, Jaroslav T. Optimization of conditions for in vitro pollen germination and tube growthin potatoes[J]. International Journal of Plant Sciences,1996,157(5):561-566.
[83] Lush W M, Clarke A E. Observation of pollen tube growth in Nicotiana alata and their implications forthe mechanism of self-incompatibility[J]. Sex Plant Reprod,1997,10:27-35.
[84] Luu D T, Qin X, Morse D, et al. S-RNase uptake by compatible pollen tubes in gametophyticself-incomp-atibility[J]. Nature,2000,407:649-651.
[85] Luu D T, Qin K, Laublin G, et al. Rejection of S-heteroallelie polen by a dualspecific S-RNase inSolanum chacoense predicts a multineric SI pollen component[J]. Genetics,2001,159(1):329-335.
[86] Mayfield J A, Fiebig A, Johnstone S E, et al. Gene families from the Arabidopsis thaliana pollen coatproteome[J]. Science,2001,292(5526):2482-2485.
[87] McClure B A, Harin V, Ebert P R, et al. Style Self-incompatibility gene products of Nicotiana alata areribonucleases[J]. Nature,1989,342:955-957.
[88] McClure B A, Gray J E, Anderson M A, et al. Self-incompatibility in Nicotiana alata involvesdegradation of pollen rRNA[J]. Nature,1990,347:757-760.
[89] Mcclura B, Mou B, Canevascini S, et al. A small asparagine-rich protein required for S-allele-specificpollen rejection in Nicotiana[J]. Proc Natl Acad Sci USA,1999,9:13548-13553.
[90] Mcclure B A, Haring V, Ebert P R, et al. Style self-incompatibility gene products of Nicotlana alata areribonucleases[J]. Nature,1989,342:955-957.
[91] Minamikawa M, Kakui H, Wang S, et al. Apple S locus region represents a large cluster of related,polymorphic and pollen-specific F-box genes[J]. Plant Mol Biol,2010,74:143-154.
[92] Murase K, Shiba H, Iwano M, et al. Membrane-anchored protein kinase involved in Brassicaself-incompalibilily signaling[J]. Science,2004,303:1516-1519.
[93] Murfett J, Atherton T L, Mou B, et al. S-RNase expressed in transgenic Nicotiana causes S-allele-specificpollen rejection[J]. Nature,1994,367:563-566.
[94] Murphy D J. The extracellularpollen coat inmembers of the Brassicaceae: composition, biosynthesis, andfunctions in pollination[J]. Protoplasma,2006,288(1-3):31-39.
[95] Muleaphy G B, Mulcaphy D L. The effect of supplemented media on the growth in vitro of bi-andtri-nucleate pollen [J]. Plant Science,1988,55(3):213-216.
[96] Nasrallah J B, Kao T H, Goldberc M L, et al. A cDNA Clone Encoding S-specific Glycoprotein FromBrassica Oleracea[J]. Nature,1985,318:263-267.
[97] Nasrallah J B, Nasrallah M E. Pollen-stigma signaling in the sporophytic self-incompatibility response[J].Plant Cell,1993,5:1325-1335.
[98] Nasrallah J B, Yu S M, Nasrallah M E. Self-incompatibility genes of Brassica olerace: expression,isolation, and structure[J]. Proc Natl Acad Sci USA,1988,85(15):5551-5555.
[99] Natthani S, Chookajom T, Ripoll D R, et al. Structural modules for receptor dimerization in the S-locusroceptor kinase extracellular domain[J]. Pans,2007,104(29):12211-12216.
[100] Nepi M, Franchi G G, Pacini E. Pollen hydration status at dispersal: cytophysiological features andstrategies[J]. Protoplasma,2001,216:171-180.
[101] Newbigin E, Vierstra R D. Sex and self denial[J]. Nature,2003,421:229.
[102] Noir S, Brautigam A, Colby T, et al. A reference map of the Arabidopsis thaliana mature pollenproteome[J]. Biochem Bioph Res Co,2005,337(4):1257-1266.
[103] O’Brien M, Kapfer C, Major G, et al. Molecular analysis of the stylar-expressed Solanum chacoensesmall asparagine-rich protein family related to the HT modifier of gametophytic self-incompatibility inNicotiana[J]. Plant J,2002,32(6):985-996.
[104] Ockendon D J. Distrbution of self-incompatibility alleles and breeding structure of openpollinatedcultivars of brussela sprouts[J]. Heredity,1974,33:159-171.
[105] Ockendon D J. An S-allele survey of cabbage (Brasdca oleracea var. capitata)[J]. Euphytica,1982,31:325-331.
[106] Omura M, Akihama T A. Pollen preservation of fruit trees for gene banks in Japan[J]. Plant GeneticResources News Letter,1980,43:28-31.
[107] Pedersen S, Simonsen V, Loeschcke V. Overlap of gametophytic and sporophytic expression in barley[J].Theor Appl Gen,1987,75:200-206.
[108] Prell H. Das Problem der Unfruchtbarkeit[J]. Nature Wochschr, N. F,1921,20:440-446.
[109] Qiao H, Wang H, Zhao L, et al. The F-box protein AhSLF-S2physically interacts with S-Rnases thatmay be inhibited by the ubiquitin/26S proteasome pathway of protein degradation during compatiblepollination in Antirrhinum[J]. Plant Cell,2004,16(3):582-595.
[110] Rajasekharan P E, Rao T M, Janakiram T, et al. Freeze preservation of gladiolus pollen[J]. Euphytica,1994,80:105-109.
[111] Ram Dixit, June B, Nasrallah. Recognizing self in the self-incompatibility response[J]. Plant Physiol,2001,125:105-108.
[112] Sanchez-Perez R, Ortega E, Henri D, et al. Inheritance and relationships of important agronomic traits inalmond[J]. Euphytica,2007,155(3):381-391.
[113] Richards A J. Plant breeding systerms[M]. Allen Unwin: London,1986.
[114] Royo J, Kunz C, Kowyama Y, et al. Loss of a histidine residue at the active site of S-locus ribonucleaseis associated with self-incompatibility in Lycopersicon peruvianum[J]. Proc Natl Acad Sci USA,1994,91(14):6511-6514.
[115] Rudd J J, Franklin-Tong V E. Unravelling response-specificity in Ca2+signalling pathways in plantcells[J]. New Phytol,2001,151(1):7-33.
[116] Sadao Komori, Junichi Soejima, Yuji Ito, et al. Analyses of the sell-incompatibilily genotype in someapple cultivars[J]. J Japan Soc Hort Sci,1998,67(6):917-926.
[117] Sage T L, Sampson F B. Evidence for ovarian self-incompatibility as a cause of self-sterility in therelictual woody angiosperm, Pseudowintera axillaris (Winteraceae)[J]. Ann Bot,2003,91(7):807-816.
[118] Sassa H, Hirano H, Ikehashi H. Identification and characterization of stylar glycoproteins associatedwith self-incompatibility genes of Japanese pear, Pyrus serotina Rehd[J]. Mol Gen Genet,1993,241(1-2):17-25.
[119] Sassa H, Nishio T, Kowyama Y, et al. Self-incompatibility (S) alleles of the Rosaceae encode membersof a distinct class of the T2/S ribonuclease superfamily[J]. Mol Gen Genet,1996,250(5):547-557.
[120] Sassa H, Hirano H, Nisho T, et al. Style-specific self-compatible mutation caused by deletion of theS-RNase gene in Japanese pear(Pyrus serontia)[J]. Plant J,1997,12(1):223-227.
[121] Sassa H, Hirano H, Ikehashi H. Self-incompatibility-related RNases in styles of Japanese pear (Pyrusserotina Rehd.)[J]. Plant Cell Physiol,1992,33(6):811-814.
[122] Sassa H, Kakui H, Minamikawa M. Pollen-expressed F-box gene family and mechanism ofS-RNase-based gametophytic self-incompatibility (GSI) in Rosaceae[J]. Sex Plant Reprod,2010,23(1):39-43.
[123] Sato Y, Kurihara A, Abe K, et al. Mode of self-compatibility inheritance in Japanese pear[J]. J JapanSoc Hort Sci,1988,57(2):76-77.
[124] Sawhney V K, Chen K, Sussex I M. Soluble proteins of the mature floral organs of tomato[J]. J PlantPhysiol,1985,121(3):265-271.
[125] Schopfer C R, Nasrallah M E, Nasrallah J B. The male determinant of self-incompatibility in Brassica[J].Science,1999,286(5445):1697-1700.
[126] Sears E R. Cytological phenomena connected with self-sterility in the flowering plants[J]. Genetics,1937,22(1),130-181.
[127] Seavey S R, Bawa K S. Late-acting self-incompatibility in angiosperms[J]. Botanical Review,1986,52(2),195-219.
[128] Shiba H, Iwano M, Entani T, et al. The dominance of alleles controlling self-incompatibility in Brassicapollen is regulated at the RNA level[J]. Plant Cell,2002,14(2):491-504.
[129] Matsumoto S, Komori S, Kitahara K, et al. S-genotype of15apple cultivars and self-compatibility of‘Megumi’[J]. J Japan Soc Hurt Sci,1999,68(2):236-241.
[130] Shimosato H, Yokota N, Shiba H. et al. Characterization of the SP11/SCR high-affinity binding siteinvolred in self/nonself recognition in Brassica self-incompatibility[J]. Plant Cell,2007,19(1):107-117.
[131] Sijacic P, Wang X, Skirpan A L, et al. Identification of the pollen determinant of S-RNase-mediatedself-incompatibility[J]. Nature,2004,29:302-305.
[132] Silva N F, Stone S L, Christie L N, et al. Expression of the S-receptor kinase in self-compatibleBrassica napus cv. Westar leads to the allele specific rejection of self-incomopatible Brassica napuspollen[J]. Mol Genetics Genomics,2001,265:552-559.
[133] Sims T L, Ordanic M. Identification of a S-ribonuclease-binding protein in Petunia hybrida[J]. PlantMol Biol,2001,47(6):771-783.
[134] Singh A, Ai Y, Kao T H. Characterization of ribonuclease activity of three S-allele-associated proteinsof Petunia inflata[J]. Plant Physiol,1991,96(1):61-68.
[135] Sparks D, Yates I E. Pecan pollen stored overa decaderetains viability[J]. HortScience,2002,37(1):176-177.
[136] Stein J C, Howlett B, Boyes D C, et al. Molecular cloning of a putative receptor protein kinase geneencoded at the self-incompatibility locus of Brassica oleracea[J]. Proc Natl Acad Sci USA,1991,88(19):8816-8820.
[137] Stone S L, Arnoldo M, Goring D R. A breakdown of Brassica seIf-incompatibility in ARC1antisensetransgenic plants[J]. Science,1999,286(5445):1729-1731.
[138] Stone S L, Anderson E M, Mullen R T, et al. ARC1is an E3ubiquitin ligase and promotes theubiquitination of proteins during the rejection of self-incompatible Brassica pollen[J]. Plant Cell,2003,15(4):885-898.
[139] Suzuki G, Kai N, Hirose T, et al. Genomic organization of the S locus: Identification andcharacterization of genes in the SLG/SRK region of S9haplotype of Brassica campestris(syn. rapa).[J].Genetics,1999,153(1):391-400.
[140] Takayama S, Shiba H, Iwano M, et al. The pollen determinant of self-incompatibility in Brassicacampestris[J]. Proc Natl Acad Sci,2000,97(4):1920-1925.
[141] Takasaki T, Hatakeyyma K, Suzuki G, et al. The S rceeptor kinase determines self-incompatibility inBrassica stigma[J]. Nature,2000,403:913-916.
[142] Tantikanjana T, Nasrallah M E, Stein J C, et al. An altemative transcript of the S locus glyeoproteingene in a class Ⅱ pollen-recessive self-incompatibility haplotype of Brassica oleracea encodes amembrane anchored protein[J]. Plant Cell,1993,5(6):657-666.
[143] Entani T, Iwano M, Shiba H, et al. Comparative analysis of the self-incompatibility (S-)locus region ofPrunus mume: identification of a polen-expressed F-box gene with allelic diversity[J]. Genes Cells,2003,8(3):203-213.
[144] Kao T H, Tsukamoto T. The molecular and genetic bases of S-RNase-based self-incompatlbility[J].Plant Cell,2004,16(Supp1.): S72-S83.
[145] Thomas S, Osman K, de Graaf B H J, et al. Investigating mechanisms involved in theself-incompatibility response in Papaver rhoeas[J]. Philos Trans R Soc Lond B,2003,358(1434):1033-1036.
[146] Ton L D, Krezdorn A H. Growth of pollen tubes in three incompatibe citrus varieties[J]. Proc Amer SocHort Sci,1966,80:211-215.
[147] Tsai D S, Lee H S, Post L C, et al. Sequence of an S-protein of Lycopersicon peruvianum andcomparison with other solanaceous S-proteins[J]. Sex Plant Reprod,1992,5(4):256-263.
[148] Tsukamoto T, Ando T, Watanabe H, et al. Duplication of the S-Locus F-box gene is associated withbreakdown of pollen function in an S-haplotype identined in a natural population of self-incompatiblePetunia axillaris[J]. Plant Mol Biol,2005,57(1):141-153.
[149] Tyers M, Jorgensen P. Proteolysis and the cell cycle: with the RING I do three destroy[J]. Curr OpinGenet Dev,2000,10(1):54-64.
[150] Ushijima K, Sassa H, Dandekar A M, et al. Structural and transcriptional analysis of theself-incompatibility locus of almond: Identification of a pollen-expressed F-box gene withhaplotype-specific polymorphism[J]. Plant Cell,2003,15(3):771-781.
[151] Valtuena F J, Rodriguez-Riano T, Espinosa F, et al. Self-sterility in two Cytisus species (Leguminosae,Papilionoideae) due to early-acting inbreeding depression[J]. Amer J Bot,2010,97(1),123-135.
[152] Wang C S, Walling L L, Eckard K J, et al. Patterns of protein accumulation in developing anthers oflilium longiflorum correiate with histological events[J]. Amer J Bot,1992,79(2):118-127.
[153] Wheeler M J, Franklin-Tong V E, Franklin F C H. The molecular and genetic basis of pollen-pistilinteractions[J]. New Phytol,2001,151(3):565-584.
[154] Williams E G, Kaul V, Rouse J L, et al. Overgrowth of pollen tubes in embryo sacs of Rhododendronfollowing interspecific pollinations[J]. Amer J Bot,1986,34,413-423.
[155] Xue Y, Carpenter R, Dickinson H G, et al. Origin of allelic diversity in Antirrhinum S locus RNases[J].Plant Cell,1996,8(5):805-814.
[156] Yamane H, Ikeda K, Hauck N R, et al. Self-incompatibility (S) locus region of the mutated S6-haplotypeof sour cherry (Prunus cerusus) contains a functional pollen S allele and a non-functional pistil Sallele[J]. J Exp Bot,2003,54(392):2431-2437.
[157] Zhang S L, Hiratsuka S. Variation in S-protein levels in styles of Japanese Pears and the expression ofself-incompatibility[J]. J Japan Soc Hort Sci,1999,68(5):911-918.
[158] Zhang S L, Hiratsuka S. Analysis of varietal differences in self-and cross-incompatibility reactions ofJapanese pear using stylar culture technique[J]. J Japan Soc Hort Sci,1999,68:373-383.
[159]艾鹏飞,罗正荣.柿品种‘禅寺丸’花粉超低温保存研究[J].华中农业大学学报,2004,23(5):563-565.
[160]白书农,谭克辉.湖北光敏核不育水稻农垦58S叶片60kD蛋白特异性及其功能的讨论[J].植物学报,1997,39(2):189-192.
[161]陈迪新,张绍铃,陶书田.沙梨花粉原位萌发与花粉管生长特性[J].南京农业大学学报,2004,27(3):34-37.
[162]陈霜莹,常永健,赵艳华,等.果树花粉的超低温保存研究[J].华北农学报,1993,8(增刊):60-64.
[163]陈腾土,杨小华,薛妙男.沙田柚花柱蛋白对花粉管生长的影响[J].广西植物,1998,18(2):160-164.
[164]陈暄,彭英,郝姗,等.茶树花粉的离体萌发研究[J].江苏农业科学,2011(6):233-235.
[165]程伟,赵志刚,郭俊杰,等.西南桦花粉低温贮藏试验初报[J].浙江林业科技,2008,27(6):49-52.
[166]丁长奎,陈其峰,夏起洲,等.营养元素与生长调节剂对枇杷花粉萌发和座果的影响[J].中国果树,1991(4):18-20.
[167]董长江,郑常文,刘丙辰,等.诸葛菜精细胞特异蛋白多肽的研究[J].植物学报,1997,39(8):712-716.
[168]杜玉虎,张绍铃,姜雪婷,等.果梅花粉离体萌发及花粉管生长特性研究[J].西北植物学报,2006,26(9):1846-1852.
[169]樊汝汶,黄金生.梓树属两个种花柱发育的细胞形态学比较研究[J].南京林产工业学院学报,1988,3:33-38.
[170]樊汝汶,吴琼美,邹惠渝.滇楸的胚胎学研究[J].南京林产工业学院学报,1980,2:67-79.
[171]冯莎莎,杜国强,师校欣,等. DNA浓度及注射时间对苹果花粉管通道法基因转化率的影响[J].中国农学通报,2007,23(4):64-66.
[172]樊新民,牛建新,孙爱新,等.榆叶梅和海棠花粉萌发率的测定及超低温保存方法[J].湖北农业科学,2004,50(6):65-66.
[173]谷瑞升,刘群录,陈雪梅,等.木本植物蛋白提取和SDS-PAGE分析方法的比较和优化[J].植物学通报,1999,16(2):171-177.
[174]郭长奎,李疆,罗淑萍,等.扁桃花粉SFB基因的鉴定和序列分析[J].经济林研究,2009,27(3):18-23.
[175]郭从俭,钱士金,王连卿,等.楸树栽培[M].北京:中国林业出版社,1988.
[176]郭尧君.蛋白质电泳实验技术[M].北京:科学出版社,2005.
[177]顾翠花,孙明哲,王守先,等.3个紫薇品种花粉离体培养与萌发研究[J].安徽农业科学,2011,39(23):14009-14011.
[178]顾红雅,陈章良.高等植物花器官的特异性基因[J].植物生理学通讯,1993,29:393-401.
[179]顾钢,苏学强,林伟杰,等.橄榄花粉生活力相关因素对育种的影响[J].福建果树,1999(3):1-4.
[180]黄永芳,吴雪辉,何美儿,等.3种油茶植物花粉贮藏及生活力的研究[J].福建林学院学报2011,31(1):56-59.
[181]华志明.植物自交不亲和分子机理研究的一些进展[J].植物生理学通讯, l999,35(1):77-82.
[182]郝明灼,彭方仁,王改萍,等.楸树人工杂交授粉试验及种实分析[J].南京林业大学学报,2008,32(5):131-134.
[183]胡适宜.植物胚胎学实验方法(五)检查花粉在柱头上萌发和花粉管在花柱中生长的制片法[J].植物学通报,1994,11:58-60.
[184]胡适宜.被子植物胚胎学[M].北京:人民教育出版社,1982,46-51.
[185]贾继文,王军辉,张金凤,等.梓树属花粉生活力的研究[J].西北植物学报,2009,29(5):0867-0873.
[186]贾文庆,刘会超,李保栓.榆叶梅花粉生活力及贮藏特性的研究[J].广东农业科学,2011,(9):39-41.
[187]姜立杰,曾家树.芸薹属植特自交不亲和性的分子机制[J]植物学通报,2001,18(4):411-417.
[188]李秉玲,尚晓倩,刘燕.芍药花粉超低温保存4年后的生活力检测[J].北京林业大学学报,2008,30(6):145-147.
[189]李继爱,姜贺飞,赵惠恩.洋水仙花粉生活力的测定方法初探[J].中国农学通报,2010,26(17):242-245.
[190]李惠敏,秦新民,覃屏生,等.超声波提取沙田柚花粉管可溶性蛋白的初步研究[J].广西师范大学学报,2004,22(1):82-86.
[191]李嘉瑞,王彩虹.杏花粉的低压保存研究[J].西北农业大学学报,1996,24(3):1-4.
[192]李俊英,陶慧,许林,等.大叶杨花粉离体萌发和贮藏条件的研究[J].上海交通大学学报,2011,29(4):83-88.
[193]李天忠,加藤直幹,奥野智旦.红星苹果花柱S-核酸酶的分离与纯化[J].农业生物技术学报,2005,13(5):568-571.
[194]李晓,张绍铃,陶书田,等.中国樱桃与甜樱桃花粉原位萌发及花粉管生长的差异[J].西北植物学报,2007,27(3):0429-0430.
[195]李叶峰,谢慧慧,刘晓红,等.牵牛花粉表面蛋白SDS-PAGE分析[J].河南农业科学,2010,1:99-102.
[196]梁建萍,韩有志,梁胜发,等.丁香花粉生活力及其贮藏性的研究[J].山西林业科技,2000,(1):10-12.
[197]粱立,徐秉芳,郑从义.紫菜薹花粉超低温保存及其原生质体分离[J].植物学报,1993,5(2):733-738.
[198]梁有旺,彭方仁,王顺才.楸树嫩枝扦插试验初报[J].林业科技开发,2006,20(l):67-69.
[199]林鸣,曹宗巽.黄瓜器官特异蛋白的研究[J].植物学报,1996,38(7):525-529.
[200]刘宝敬,王晓佳.植物孢子体型自交不亲和性研究进展[J].生命科学,1996,8(4):38-41.
[201]刘宝敬,宋明,李成琼,等.等电聚焦电泳测定甘蓝自交不亲和性研究[J].西南农业大学学报,1998,20(2):104-110.
[202]刘会超,李继爱,姜贺飞,等.洋水仙花粉生活力的测定方法初探[J].中国农学通报,2010,26(17):242-245.
[203]刘剑锋,阎秀峰,程云清,等.高山红景天花粉的超低温保存[J].南京林业大学学报,2006,30(6):138-140.
[204]刘武林.花粉的采集、贮藏和生活力的测定[J].植物学通报,1985,3(3):8-12.
[205]刘晓红,彭丽媛,陆小平,等.牵牛花粉表面物质的释放[J].北方园艺,2006(5):124-125.
[206]刘燕,张亚利.梅花花粉超低温保存研究[J].北京林业大学学报,2004,(3):22-25.
[207]刘自刚.不同贮藏温度对桔梗花粉生活力及授粉结实能力的影响[J].热带作物学报,2011,32(5):956-959.
[208]律春燕,王雁,朱向涛,等.黄牡丹花粉生活力测定方法的比较研究[J].林业科学研究,2010,23(2):272-277.
[209]芦岩,李玉花.芸苔属中自交不亲和反应的信号转导[J].植物生理学通讯,2005,41(4):547-552.
[210]马建伟,王军辉,张守攻,等.云杉花粉的贮藏及萌发研究[J].林业科学研究,2012,25(3):302-307.
[211]毛洪玉,崔文山,王亚斌.地被菊花粉萌发及花粉活力的研究[J].辽宁林业科技,2004,(5):12-14
[212]莫花浓.沙田柚离体花粉管的培养研究[J].玉林师范学院学报,2006,27(5):83-85.
[213]孟金陵,刘定富,罗鹏,等.植物生殖遗传学[M].北京:科学出版社,1995.
[214]南矗,郑莲香,周立东.楸树的研究现状与开发前景[J].西北药学杂志,2007,22(2):90-93.
[215]牛俊海,鲁晓民,汤继华,等.禾本科植物自交不亲和性及其分子生物学研究进展[J].分子植物育种,2006,4(2):269-274.
[216]欧阳英,李秉玲,刘燕.密花石斛花粉的保存方法研究[J].北京林业大学学报,2010,32(6):151-154.
[217]齐国辉.鸭梨自交不亲和与亲和变异的生理生化特性及分子机理研究[D].保定:河北农业大学,2005.
[218]秦新民,李惠敏,薛妙男,等.沙田柚自交、异交花粉管蛋白双向电泳分析[J].广西植物,2004,24(6):566-569.
[219]秦新民,吕建珍,薛妙男,等.沙田柚和酸柚花粉壁蛋白的双向电泳分析[J].广西师范大学学报,2004,22(4):70-73.
[220]秦新民,吕建珍,李惠敏,等.沙田柚和酸柚花粉壁蛋白的双向电泳分析[J].广西师范大学学报,2004,22(4):83-87.
[221]饶桂荣,杨继华,薛妙男.沙田柚各器官特异蛋白质的研究[J].广西师范大学学报,2000,18(2):78-81.
[222]汤建,朱广廉,曹宗巽.兰州百合精细胞特异蛋白的研究[J].植物学报,1997,39(6):511-516.
[223]汤青林,宋明,王小佳.芸苔属植特自交不亲和性及其机理研究进展[J].生物工程进展,2001,2l(4):22-25.
[224]唐荣平,杨永红,苏汉林,等.丽江山慈菇花粉生活力研究[J].西南农业学报,2009,22(1):153-155.
[225]唐玲,姜卫兵,翁忙玲.楸树的园林特性及其开发应用[J].中国农学通讯,2007,23(4):276-280.
[226]王爱华,戴洪义.果树自交不亲和性的研究进展[J].莱阳农学院学报,2002,19(3):206-209.
[227]王爱云,李栒,胡大有.诸葛菜与芸薹属间花粉与柱头相互作用的研究[J].湖南农业大学学报,2006,32(3):232-236.
[228]王改萍,彭方仁,李生平.银杏叶片蛋白质含量动态变化的电泳分析[J].南京林业大学学报,2006,30(4):114-118.
[229]王改萍,徐涛,彭方仁.木本植物花粉的保存研究进展[J].林业科技开发,2007,21(6):9-11.
[230]王改萍,岑显超,何力,等.水分胁迫对楸树苗木光合特性的影响[J].南京林业大学学报,2007,31(6):57-60.
[231]王家福,刘月学,刘小军,等.枇杷花粉干冻法超低温保存研究[J].中国农学通报,2004,20(1):1-2.
[232]王君晖,黄纯农.木本植物种质超低温保存的研究进展[J].世界林业研究,1998,5:6-11.
[233]王茂广.孢子体型自交不亲和反应臂重复蛋白ARCI[J].生命科学,2007,19(1):87-89.
[234]王钦丽,卢龙斗,吴小琴,等.花粉的保存及其生活力测定[J].植物学通报,2002,19(3):365-373.
[235]王郁,江昌俊,张和禹.茶树自交花粉管在活体花柱中的不亲和性观察[J].茶叶科学,2008,28(6):429-435.
[236]王郁民,李嘉瑞.猕猴桃花粉的有机溶剂保存[J].落叶果树,1992,2:3-7.
[237]王振宇,杨春瑜,金钟跃.超声波提取芦荟凝胶的工艺[J].东北林业大学学报,2002,30(4):72-73.
[238]王铮敏.超声波在植物有效成分提取中的应用[J].三明高等专科学校学报,2002,19(4):45-53.
[239]韦仲新,王红,金巧军.中国紫藏科花粉形态的研究[J].云南植物研究,2001,23(2):225-230.
[240]吴厚久,陈竹生,胨克玲.两个甜橙选择系自变异变的亲和性观察[J].园艺学报,1986,13(4):281-282.
[241]吴华清,张绍铃,李晓,等.植物自交不亲和性的分子生物学进展[J].南京农业大学学报,2006,29(4):119-126.
[242]吴华清,衡伟,李晓,等.大果黄花梨自交亲和性变异机制研究[J].南京农业大学学报,2007,30(2):29-33.
[243]吴俊,谷超,张绍铃,等.中国杏自交不亲和花粉特异SFB基因的鉴定与序列分析.园艺学报,2010,37(8):1329-1338.
[244]武冲,张勇,仲崇禄,等.培养条件及贮藏温度和时间对木麻黄花粉萌发率的影响[J].热带亚热带植物学报,2010,18(3):316-320.
[245]徐义流,张绍铃.花粉-雌蕊相互作用的分子基础口[J].西北植物学报,2003,23(10):1800-1809.
[246]徐义流,张绍铃.梨配子体型自交不亲和性及其分子机理[J].果树学报,2003,20(1):59-63.
[247]许桂芳,张朝阳.红继木的花器构造与花粉生活力的研究[J].吉林农业大学学报,2003,25(5):520-522.
[248]许珂,古松,江莎.金银忍冬花粉离体萌发初探[J].热带亚热带植物学报,2008,16(2):109-115.
[249]薛妙男,陈腾土,杨继华.沙田柚自交和异交亲和性观察[J].园艺学报,1995,22(2):127-132.
[250]薛妙男,李义平,张杏辉,等.沙田柚自交花柱S1-RNase的免疫胶体金定位[J].广西师范大学学报,2000,18(1):81-84.
[251]薛妙男,杨继华.沙田柚花粉管在花柱中的生长途径及其识别[J].广西师范大学学报,2001,19(2):60-66.
[252]薛勇彪,崔海洋,赖钊.不亲和性和自交亲和性及其分子生物学基础[M].北京:科学出版社,2002,128-148.
[253]薛勇彪,孟金陵.高等植物自交不亲和性的分子生物学[J].生物工程进展,1995,15(1):32-42.
[254]岩波洋造.花粉学[M].东京:讲谈株式会社,1980.
[255]严伟,李淑芬,田松江.超声波协助提取技术[J].化工进展,2002,21(9):649-651.
[256]杨继华,李红艳,薛妙男.沙田柚花柱S-糖蛋白的分离与鉴定[J].广西师范大学学报,2000,18(4):66-70.
[257]杨继华,饶桂荣,薛妙男.沙田柚花柱S-糖蛋白的纯化和N-端序列测定[J].广西师范大学学报,2001,19(1):72-79.
[258]杨继华,颜承,薛妙男,等.沙田柚花粉蛋白质双向电泳分析[J].广西师范大学学报,2002,20(2):73-77.
[259]杨美珠,陈右人.茶树自交不亲和现象之观察[J].中国园艺,2000,46:83-92.
[260]杨玉珍,王顺财,彭方仁.我国揪树研究现状及开发利用策略[J].林业科技开发,2006,20(3):4-7.
[261]杨鑫,张秀省,穆红梅,等.4种槐树花粉生活力测定方法的比较[J].广东农业科学,2011,12:53-55.
[262]叶培忠,刘玉莲.促进楸树结实的研究[J].南京林产工业学院学报,1980,1:116-121.
[263]于澄字,胡胜武,郭蔼光.芸苔属植物S受体蛋白激酶的研究[J].生物技术,2002,12(1):44-45.
[264]于晓敏,蓝兴国,李玉花.泛素/26S蛋白酶体途径与显花植物自交不亲和反应[J].植物学通报,2006,23(2):197-206.
[265]袁忠海,吴道澄,李晓晔.超声波对魔芋提取葡甘露聚糖的影响[J].第四军医大学学报,2003,24(3):238-242.
[266]曾广娟,李春敏,张新忠,等.苹果实生树阶段转变特异蛋白质的SDS-PAGE分析[J].园艺学报,2008,35(7):1059-1064.
[267]赵长星,刘成连.培养基种类及蔗糖浓度对部分果树花粉发芽率的影响[J].河北林果研究,2001,16(3):240-243.
[268]张丞,王冠,易素华,等.拟南芥花粉表面蛋白质数据的整合[J].上海师范大学学报,2009,38(5):511-515.
[269]张锦.“材”貌绝伦的揪树[J].森林与人类,2003,(3):34-36.
[270]张锦,罗宁,李同顺,等.楸树播种育苗技术[J].林业科技开发,2007,21(5):83-85.
[271]张绍铃,陈迪新,康琅,等.培养基组分及pH值对梨花粉萌发和花粉管生长的影响[J].西北植物学报,2005,25(2):225-230.
[272]张绍铃,房经贵,杨记磙.果树自交不亲和性的遗传与生理机制及其研究[J].果树学报,2001,18(1):49-52.
[273]张绍铃,平塜伸,徐国华,等.梨自交不亲和及其亲和突变品种花柱内S4(S4SM)基因的表达与作用的比较[J].植物学报,2001,43(11):1172-1178.
[274]张绍铃,杨记磙,李秀根,等.梨自交不亲和强度不同品种花柱S糖蛋白含量的差异[J].园艺学报,2002,29(2):165-167.
[275]张绍铃,周建涛,徐义流,等.梨花柱半离体培养法及品种自交不亲和基因型鉴定[J].园艺学报,2003,30(6):703-706.
[276]张绍铃,平塜伸.梨花柱S糖蛋白对离体花粉萌发及花粉管生长的影响[J].园艺学报,2000,27(4):251-256.
[277]植物教研组,江苏省珍贵用材树种-楸树属的研究[J].南林科技,1977.
[278]周楠楠,方炎明,马成涛.红桤木花粉生活力及其贮藏方法的研究[J].南京林业大学学报,2010,34(5):34-38.
[279]周俊国,扈惠灵,张慧蓉,等.苹果和梨自交不亲和程度的荧光显微观察[J].河南科技学院学报,2010,38(2):27-31.
[280]张玉进,张兴国,刘佩瑛.魔芋花粉的低温和超低温保存[J].园艺学报,2000,27(2):139-140.
[281]朱开开,张春芳,杨锋,等.苹果花粉F-box基因MdSL FB9克隆与鉴定[J].西北植物学报,2009,29(4):0650-0655.