抗寒苹果、梨种质资源遗传多样性研究
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摘要
本文对国家果树种质公主岭寒地果树圃保存的抗寒苹果292、梨210份资源进行了表型性状——树姿、树型、树势、一年生枝、叶片、花、果实等基本信息进行描述，同时对这些资源进行了生物学特性——物候期、萌芽率、成枝力、丰产性、矮化性、果实品质、抗寒性、抗病性、染色体倍性等进行鉴定、评价，得到极其丰富的表型遗传多样性。在此基础上，首次建立了抗寒苹果、梨种质资源数据库，提交给国家科技平台建设——E平台，为资源利用提供更广泛的共享。本文应用RAPD技术首次对上述资源进行了分子标记，对其表型遗传多样性进行检测，从分子层面揭示了抗寒苹果、梨的遗传多样性。在此基础上，又结合50余年抗寒苹果、梨育种经验，首次构建了抗寒苹果、梨核心种质。同时应用RAPD分子标记技术首次获得了抗寒苹果、梨种质特异标记。分析研究结果，提出将保存的抗寒苹果资源中的杂交群体分类归属上，应在经典分类学中的真正苹果组中单独列为一系——杂交群系；山楂海棠与陇东海棠在聚类图中，表现出较远的亲缘关系，与山定子亲缘关系较近。实际上山楂海棠和山定子与陇东海棠与陇东海棠野生原始分布也较远。山楂海棠和山定子原产于长白山区，而陇东海棠原产于甘肃，二者有远缘的地理关系。苹果梨、库尔勒香梨因其在聚类图上与白梨聚在一起，表现出与白梨有较近的亲缘关系，因此，将二者在分类学上划分在白梨系统。
上述研究为抗寒苹果、梨资源深入研究、加速育种进程——亲本选择、早期鉴定等工作提供了科学依据。主要结果如下：
1．本研究对500余份抗寒苹果、梨种质资源的树姿、树型、树势、一年生枝、叶片、花、果实等植物学基本性状进行了较为详细的调查，和对物候期、萌芽率、成枝力、丰产性、矮化性、果实品质、抗寒性、抗病性、染色体倍性等生物学特性进行鉴定、评价的系统研究，掌握了这些资源大量的基本信息：从高大乔木到矮化；从小果型到大果型；从果皮黄白到浓红色；从抱合直立生树姿到开张下垂；果实风味从酸涩苦到甜；从产量极低到丰产；从极抗寒到不抗寒；从极抗病到不抗病，等等这些性状，表现出抗寒苹果、梨种质资源极为丰富的遗传多样性。
这些研究为抗寒苹果、梨种质资源数据库建立及初级核心种质构建提供了依据。
2．本研究对抗寒苹果、梨种质资源果实中可溶性固形物、糖、酸和维生素C、进行了分析测定。可溶性固形物含量、糖、酸和维生素C含量也表现出丰富的多样性。总的看，苹果资源中小果型果实的可溶性固形物含量、糖、酸和维生素C含量均高于大果型果实，也就是说，山定子高于海棠果高于杂交后代高于大苹果。
3．本研究对抗寒苹果、梨种质资源抗寒性鉴定、评价，结果是：苹果资源抗寒能力最强的是山定子(M．baccata Borkh)、毛山定子(M．mandshurica(Maxim．)Kom．)山楂海棠(M．komarovii(sarg)Rehd．)、小酸果等及含有山定子或毛山定子血缘的海棠果(M．prunifolia Brokh．)类；含有苹果(M．pumila)或塞威氏苹果(M．sieversii)血缘的大苹果类抗寒能力弱(如绿香蕉、丹苹一号)；而海棠果与苹果(M．pumila)类的杂种后代趋中变异(如金红、龙冠)，其抗寒能力介于两者之间，并呈连续性正态分布。梨资源抗寒能力强的是秋子梨(P．ussuriensis Maxim．)、白梨(P．bretschneideri Rehd．)次之；洋梨(P．communis L．)抗寒力最差。而秋子梨(P．ussuriensis Maxim．)与白梨(P．bretschneideri Rehd．)、洋梨(P．communis L．)的种间杂种后代趋中变异，平均抗寒能力介于两者之间。同时也有个别后代是超亲遗传。抗寒能力最强的可抗—45℃以上的极端低温。
4．本研究对抗寒苹果、梨种质资源抗病性鉴定、评价是：苹果资源抗苹果腐烂病能力最强的是山楂海棠、扎矮山定子、小黄海棠、黄太平、小酸果、大秋等；梨资源抗梨黑星病能力最强的是秋子梨中的酸梨锅子、真红宵梨和西丰京白，他们同时又是抗寒的优良资源。本研究还得出抗寒苹果资源材料抗寒力与抗苹果腐烂病能力二者之间呈正相关，既抗寒能力强的资源材料，其抗苹果腐烂病能力也越强。这对抗寒苹果、梨种质资源的深入研究、构建核心种质和育种利用提供了科学依据和优异的资源。
5．本研究对抗寒苹果、梨种质资源染色体倍数性鉴定结果是，所有的供试资源不论分类学上属于哪个种，不论果实、叶片大小均是二倍体，2n=2x=34。
6．本研究首次在对抗寒苹果、梨种质资源表型性状描述、鉴定评价研究基础上，建立了抗寒苹果、梨种质资源数据库。
7．本研究首次对抗寒苹果、梨种质资源进行RAPD分子标记研究，构建抗寒苹果、梨RAPD反应体系及优化反应程序，筛选出25个引物。用于对供试的266份苹果、107份梨进行RAPD随机扩增，RAPD扩增带的多态性百分率分别为93.47％、96.2％。使用Nei-Li遗传距离，对180份苹果、87份梨资源的RAPD指纹图谱进行分析研究，并计算资源间的遗传距离。其范围苹果在0.08911～1.0000之间，平均0.00506、金红芽变4号与金红芽变7号的遗传距离最小，仅为0.08911；陇东海棠与铃铛果遗传距离最大，为1.0000。梨在0.0400～0.4269之间，平均0.1934。酸梨锅子与龙香梨的遗传距离最小，仅为0.0400；黑龙江红花盖与远东一号遗传距离最大，为0.4269。
8．本研究对聚类图进行分析，得出苹果资源中的山定子系、海棠系和陇东海棠系有着相对较近的亲缘关系，这与经典分类系大体一致。然而，供试的苹果属资源材料的表型性状遗传多样性非常丰富。山定子与海棠果表型性状相近，在此可以断定二者亲缘关系最近。同时，也进一步证明前人所做的结论——海棠果是山定子与大苹果的杂交后代。但是，陇东海棠、山楂海棠的表型性状与山定子有着较大差异。前两者叶片与山楂叶片相似。那么，结合聚类图分析，作者推断，陇东海棠、山楂海棠可能是山定子与同属蔷薇科中的山楂远缘杂交后代，因苹果属、梨属、山楂属、花楸属具有共祖关系。得出梨资源材料的表型性状遗传多样性亦非常丰富。其中的苹果梨、库尔勒香梨因其在聚类图上与白梨聚在一起，表现出与白梨有较近的亲缘关系，因此，将二者在分类学上划分在白梨系统。
9．本研究首次对抗寒苹果杂交后代资源材料分类归属问题结论是，依照经典分类学，将杂交后代统一划分为杂交群系，在真正苹果组中单独列出一系。
10．本研究首次依据对抗寒苹果、梨种质资源植物学、生物学特征特性进行描述及对其抗性鉴定评价、RAPD分子标记分析研究结果，提出并构建了抗寒苹果、梨种质资源的核心种质，苹果46份资源，梨29份资源。
11．本研究首次获得抗寒苹果、梨种质资源RAPD特异标记。
In the present dissertation, 292 winterhardy apple accessions and 210winterhardy pear accessions conserved in the national fruit germplasm Gongzhulinghardy fruit nursery were employed as the experimental materials to describe theirphenotypic characters including tree posture, tree type and characteristics ofone-year-old branch, leaf, flower and fruit; to evaluate and determine their biologiccharacteristics including phenological period (phenophase), sprouting rate, branchingability, cropping efficiency, dwarfing ability, fruit quality, cold-resistance,disease-resistance and chromosome ploidy; so that the phenotypic genetic diversitywas found.
On the base of phenotypic results, the database about winterhardy apple and peargermplasm resources was set up and submitted to E-deck constructed by the nationalscience and technology ministry in order to provide the broad share and utilization.
RAPD technique was used to make molecular marker in all experimentalmaterials so that the phenotypic genetic diversity was screened and verified, then thegenetic diversity of winterhardy apple and pear was further revealed in molecularlevel. Combining the result in molecular marker and the experience accumulatedduring the past 50 years in winterhardy apple and pear breeding, the core collectionsin winterhardy apple and pear were first constructed and the particular RAPDmolecular markers were gained from winterhardy apple and pear germplasmresources.
By the further analysis of the research results, we proposed that the hybridprogeny (group) in winterhardy apple resources maintained at the repository wasclassified by itself and formed a new serial -- hybrid group serial in Sect. Malus; onthe cluster graph, M. komarovii had long distance with M. kansuensis illustrated thefar genetic relationship, and short distance with M. baccata illustrated the closegenetic relationship. In fact, both M. komarovii and M. baccata originated atChangbai mountain area, but M. kansuensis originated at Gansu province, so thegeographic distance between two origin places is very big. Because Pingguoli andKuerlexiangli were clustered together with Pyrus bretschneideri, so these varietiesillustrated the near relationship with P. bretschneideri, then were assorted into thesystem of P. bretschneideri.
By the all research results above, the scientific foundation was made for furthergermplasm resource research, speedup of breeding procedure, parent selection, early stage identification and so on. Main results were as follows:
1. The basic information of 502 winterhardy apple and pear accessions wascollected by the detailed investigation of botanic characters including tree posture,tree type and characteristics of 1-year-old branch, leaf, flower and fruit; and theevaluation and determination of their biologic characteristics including phenologicalperiod (phenophase), sprouting rate, branching ability, cropping efficiency, dwarfingability, fruit quality, cold-resistance (hardiness), disease-resistance and chromosomeploidy. Then, substantial genetic diversity was found in many (phenotypic) characters;i. e. from tall arbor to bush, from big to small fruit, from yellowish to strong redpericarp, from upright to drooping tree posture, from sour, astringent, bitter to sweetfruit flavor, from low to high productivity, from strong to weak cold-resistance, fromstrong to weak disease-resistance and so on. All of these results made it possible to setup the database and to construct the preliminary core collections among winterhardyapple and pear germplasm resources.
2. By the analysis and measuration of soluble solid content, sugar content, acidcontent and vitamin C content in the fruit of winterhardy apple and pear germplasmresources, the genetic diversity was verified in the characters above. General speaking,the content of soluble solid, sugar, acid and Vitamin C of the small and middle-sizefruit in apple resources were higher than those of big-size fruit, that is, the order fromhigh to low content in fruit was M. baccata > M. asatica > hybrid progeny > M.domestica.
3. By the evaluation and determination of cold-resistance about winterhardy appleand pear germplasm resources, the apple resources of strong cold resistance were M.baccata Borkh., M. mandshurica (Maxim.) Kom., M. komarovii (sarg) Rehd.,Xiaosuanguo and M. prunifolia Borkh. which had the genetic relationship with M.baccata Borkh. or M. mandshurica (Maxim.) Kom., but apple resources with big-sizefruit e. g. Lvxiangjiao, Danpingyihao, which had the genetic relationship with M.domestica or M. sieversii, had the weak cold resistance; the hybrid progeny betweenM. prunifolia Borkh. and M. domestica e. g. Jinhong, Longguan, had the moderatecold resistance and displayed the normal distribution. In the winterhardy pearresources, P. ussuriensis Maxim. had the strong cold resistance, then P. bretschneideriRehd. middle; and then P. communis L. had the weak cold resistance. The hybridbetween P. ussuriensis Maxim. and P. bretschneideri Rehd. or P. communis L. had themoderate cold resistance and displayed the central tendency variation, but some individuals displayed super-parent heredity. The resources with the strongest coldresistance could resist minus 45℃.
4. By the evaluation and determination of disease-resistance about winterhardyapple and pear resources, the resources with the strong resistance to apple canker wereM. komarovii, Zha'aishandingzi, Xiaohuanghaitang, Huangtaiping, Xiaosuanguo andDaqiu; the resources with the strong resistance to pear scab were Suanliguozi,Zhenhongxiao and Xifengjingbai, which were very good winterhardy pear germplasmresources. It was also found that the resources which had good cold resistance alsohad the good disease resistance; there was the positive correlation between thecharacters. These results could make a good scientific base for further research,construction of core collections and utilization in breeding about winterhardy appleand pear germplasm resources.
5. By the determination of chromosome ploidy on winterhardy apple and peargermplasm resources, all of the experimental materials belonged to diploid 2n=2x=34, no matter species, fruit or leaf.
6. Based on the results of phenotypic character description, evaluation anddetermination, the database was set up about winterhardy apple and pear germplasmresources.
7. Using the winterhardy apple and pear germplasm resources, RAPD reactionsystem and optimizing reaction procedure were established, 25 primer were selectedand used in 266 apple accessions and 107 pear accessions. The percentage of thepolymorphism bands by RAPD amplifying was 93.47% and 96.20% for apple andpear respectively. By the Nei-Li genetic distance, 180 apple accessions and 87 pearaccessions were analysis and calculated the value of the genetic distance, which was0.0891 to 1.000 in apple (average value 0.00506) and 0.040 to 0.4269 in pear (averagevalue 0.1394). In apple resources, Jinhong4hao and Jinhong7hao had the smallestgenetic distance 0.08911, but M. kansuensis and LingDangGuo had the biggestgenetic distance 1.0000. In pear resources, Suanliguozi and Longliangli had theminimum genetic distance 0.040, however, Heilongjianghonghuagai andYuandongyihao had the maximum genetic distance 0.4269.
8. By the analysis of cluster graph, it was found that Sect. M. baccata, Sect. M.prunifolia and Sect. M. kansuensis had the close genetic relationship which was samewith the classic taxology. But the phenotypic genetic diversity was very rich(abundant) in the Malus resources. Because the phenotypic character between M. baccata and M. prunifolia was close, they had the nearest genetic relationship. It wasverified that M. prunifolia was the hybrid between M. baccata and M. domestica,which was reported before. But there were obvious differences between M.kansuensis or M. komarovii and M. baccata in the phenotypic characters. Combinedwith the analysis of the cluster graph, the present author proposed that M. kansuensisand M. komarovii were derived from the cross of Malus and Crataegus because Malus,Pyrus, Crataegus and Sorbus had the same ancient species. About pear, there wasalso the abundant phenotypic diversity. Because Pingguoli and Kuerlexiangli wereclustered together with P. bretschneideri, and illustrated the near genetic relationshipwith P. bretschneideri, so they were assorted into the system of P. bretschneideri.
9. According to the classic taxology, hybrid progeny in winterhardy appleresources was formed a new series -- hybrid group series in Sect. Malus.
10. According to the results in botanic and biologic investigation, evaluation anddetermination, and the result in RAPD molecular marker, core collections inwinterhardy apple and pear germplasm resources was constructed, which were 46accessions in apple and 29 accessions in pear.
11. Particular RAPD markers were gained from winterhardy apple and peargermplasm resources.
上述研究为抗寒苹果、梨资源深入研究、加速育种进程——亲本选择、早期鉴定等工作提供了科学依据。主要结果如下：
1．本研究对500余份抗寒苹果、梨种质资源的树姿、树型、树势、一年生枝、叶片、花、果实等植物学基本性状进行了较为详细的调查，和对物候期、萌芽率、成枝力、丰产性、矮化性、果实品质、抗寒性、抗病性、染色体倍性等生物学特性进行鉴定、评价的系统研究，掌握了这些资源大量的基本信息：从高大乔木到矮化；从小果型到大果型；从果皮黄白到浓红色；从抱合直立生树姿到开张下垂；果实风味从酸涩苦到甜；从产量极低到丰产；从极抗寒到不抗寒；从极抗病到不抗病，等等这些性状，表现出抗寒苹果、梨种质资源极为丰富的遗传多样性。
这些研究为抗寒苹果、梨种质资源数据库建立及初级核心种质构建提供了依据。
2．本研究对抗寒苹果、梨种质资源果实中可溶性固形物、糖、酸和维生素C、进行了分析测定。可溶性固形物含量、糖、酸和维生素C含量也表现出丰富的多样性。总的看，苹果资源中小果型果实的可溶性固形物含量、糖、酸和维生素C含量均高于大果型果实，也就是说，山定子高于海棠果高于杂交后代高于大苹果。
3．本研究对抗寒苹果、梨种质资源抗寒性鉴定、评价，结果是：苹果资源抗寒能力最强的是山定子(M．baccata Borkh)、毛山定子(M．mandshurica(Maxim．)Kom．)山楂海棠(M．komarovii(sarg)Rehd．)、小酸果等及含有山定子或毛山定子血缘的海棠果(M．prunifolia Brokh．)类；含有苹果(M．pumila)或塞威氏苹果(M．sieversii)血缘的大苹果类抗寒能力弱(如绿香蕉、丹苹一号)；而海棠果与苹果(M．pumila)类的杂种后代趋中变异(如金红、龙冠)，其抗寒能力介于两者之间，并呈连续性正态分布。梨资源抗寒能力强的是秋子梨(P．ussuriensis Maxim．)、白梨(P．bretschneideri Rehd．)次之；洋梨(P．communis L．)抗寒力最差。而秋子梨(P．ussuriensis Maxim．)与白梨(P．bretschneideri Rehd．)、洋梨(P．communis L．)的种间杂种后代趋中变异，平均抗寒能力介于两者之间。同时也有个别后代是超亲遗传。抗寒能力最强的可抗—45℃以上的极端低温。
4．本研究对抗寒苹果、梨种质资源抗病性鉴定、评价是：苹果资源抗苹果腐烂病能力最强的是山楂海棠、扎矮山定子、小黄海棠、黄太平、小酸果、大秋等；梨资源抗梨黑星病能力最强的是秋子梨中的酸梨锅子、真红宵梨和西丰京白，他们同时又是抗寒的优良资源。本研究还得出抗寒苹果资源材料抗寒力与抗苹果腐烂病能力二者之间呈正相关，既抗寒能力强的资源材料，其抗苹果腐烂病能力也越强。这对抗寒苹果、梨种质资源的深入研究、构建核心种质和育种利用提供了科学依据和优异的资源。
5．本研究对抗寒苹果、梨种质资源染色体倍数性鉴定结果是，所有的供试资源不论分类学上属于哪个种，不论果实、叶片大小均是二倍体，2n=2x=34。
6．本研究首次在对抗寒苹果、梨种质资源表型性状描述、鉴定评价研究基础上，建立了抗寒苹果、梨种质资源数据库。
7．本研究首次对抗寒苹果、梨种质资源进行RAPD分子标记研究，构建抗寒苹果、梨RAPD反应体系及优化反应程序，筛选出25个引物。用于对供试的266份苹果、107份梨进行RAPD随机扩增，RAPD扩增带的多态性百分率分别为93.47％、96.2％。使用Nei-Li遗传距离，对180份苹果、87份梨资源的RAPD指纹图谱进行分析研究，并计算资源间的遗传距离。其范围苹果在0.08911～1.0000之间，平均0.00506、金红芽变4号与金红芽变7号的遗传距离最小，仅为0.08911；陇东海棠与铃铛果遗传距离最大，为1.0000。梨在0.0400～0.4269之间，平均0.1934。酸梨锅子与龙香梨的遗传距离最小，仅为0.0400；黑龙江红花盖与远东一号遗传距离最大，为0.4269。
8．本研究对聚类图进行分析，得出苹果资源中的山定子系、海棠系和陇东海棠系有着相对较近的亲缘关系，这与经典分类系大体一致。然而，供试的苹果属资源材料的表型性状遗传多样性非常丰富。山定子与海棠果表型性状相近，在此可以断定二者亲缘关系最近。同时，也进一步证明前人所做的结论——海棠果是山定子与大苹果的杂交后代。但是，陇东海棠、山楂海棠的表型性状与山定子有着较大差异。前两者叶片与山楂叶片相似。那么，结合聚类图分析，作者推断，陇东海棠、山楂海棠可能是山定子与同属蔷薇科中的山楂远缘杂交后代，因苹果属、梨属、山楂属、花楸属具有共祖关系。得出梨资源材料的表型性状遗传多样性亦非常丰富。其中的苹果梨、库尔勒香梨因其在聚类图上与白梨聚在一起，表现出与白梨有较近的亲缘关系，因此，将二者在分类学上划分在白梨系统。
9．本研究首次对抗寒苹果杂交后代资源材料分类归属问题结论是，依照经典分类学，将杂交后代统一划分为杂交群系，在真正苹果组中单独列出一系。
10．本研究首次依据对抗寒苹果、梨种质资源植物学、生物学特征特性进行描述及对其抗性鉴定评价、RAPD分子标记分析研究结果，提出并构建了抗寒苹果、梨种质资源的核心种质，苹果46份资源，梨29份资源。
11．本研究首次获得抗寒苹果、梨种质资源RAPD特异标记。
In the present dissertation, 292 winterhardy apple accessions and 210winterhardy pear accessions conserved in the national fruit germplasm Gongzhulinghardy fruit nursery were employed as the experimental materials to describe theirphenotypic characters including tree posture, tree type and characteristics ofone-year-old branch, leaf, flower and fruit; to evaluate and determine their biologiccharacteristics including phenological period (phenophase), sprouting rate, branchingability, cropping efficiency, dwarfing ability, fruit quality, cold-resistance,disease-resistance and chromosome ploidy; so that the phenotypic genetic diversitywas found.
On the base of phenotypic results, the database about winterhardy apple and peargermplasm resources was set up and submitted to E-deck constructed by the nationalscience and technology ministry in order to provide the broad share and utilization.
RAPD technique was used to make molecular marker in all experimentalmaterials so that the phenotypic genetic diversity was screened and verified, then thegenetic diversity of winterhardy apple and pear was further revealed in molecularlevel. Combining the result in molecular marker and the experience accumulatedduring the past 50 years in winterhardy apple and pear breeding, the core collectionsin winterhardy apple and pear were first constructed and the particular RAPDmolecular markers were gained from winterhardy apple and pear germplasmresources.
By the further analysis of the research results, we proposed that the hybridprogeny (group) in winterhardy apple resources maintained at the repository wasclassified by itself and formed a new serial -- hybrid group serial in Sect. Malus; onthe cluster graph, M. komarovii had long distance with M. kansuensis illustrated thefar genetic relationship, and short distance with M. baccata illustrated the closegenetic relationship. In fact, both M. komarovii and M. baccata originated atChangbai mountain area, but M. kansuensis originated at Gansu province, so thegeographic distance between two origin places is very big. Because Pingguoli andKuerlexiangli were clustered together with Pyrus bretschneideri, so these varietiesillustrated the near relationship with P. bretschneideri, then were assorted into thesystem of P. bretschneideri.
By the all research results above, the scientific foundation was made for furthergermplasm resource research, speedup of breeding procedure, parent selection, early stage identification and so on. Main results were as follows:
1. The basic information of 502 winterhardy apple and pear accessions wascollected by the detailed investigation of botanic characters including tree posture,tree type and characteristics of 1-year-old branch, leaf, flower and fruit; and theevaluation and determination of their biologic characteristics including phenologicalperiod (phenophase), sprouting rate, branching ability, cropping efficiency, dwarfingability, fruit quality, cold-resistance (hardiness), disease-resistance and chromosomeploidy. Then, substantial genetic diversity was found in many (phenotypic) characters;i. e. from tall arbor to bush, from big to small fruit, from yellowish to strong redpericarp, from upright to drooping tree posture, from sour, astringent, bitter to sweetfruit flavor, from low to high productivity, from strong to weak cold-resistance, fromstrong to weak disease-resistance and so on. All of these results made it possible to setup the database and to construct the preliminary core collections among winterhardyapple and pear germplasm resources.
2. By the analysis and measuration of soluble solid content, sugar content, acidcontent and vitamin C content in the fruit of winterhardy apple and pear germplasmresources, the genetic diversity was verified in the characters above. General speaking,the content of soluble solid, sugar, acid and Vitamin C of the small and middle-sizefruit in apple resources were higher than those of big-size fruit, that is, the order fromhigh to low content in fruit was M. baccata > M. asatica > hybrid progeny > M.domestica.
3. By the evaluation and determination of cold-resistance about winterhardy appleand pear germplasm resources, the apple resources of strong cold resistance were M.baccata Borkh., M. mandshurica (Maxim.) Kom., M. komarovii (sarg) Rehd.,Xiaosuanguo and M. prunifolia Borkh. which had the genetic relationship with M.baccata Borkh. or M. mandshurica (Maxim.) Kom., but apple resources with big-sizefruit e. g. Lvxiangjiao, Danpingyihao, which had the genetic relationship with M.domestica or M. sieversii, had the weak cold resistance; the hybrid progeny betweenM. prunifolia Borkh. and M. domestica e. g. Jinhong, Longguan, had the moderatecold resistance and displayed the normal distribution. In the winterhardy pearresources, P. ussuriensis Maxim. had the strong cold resistance, then P. bretschneideriRehd. middle; and then P. communis L. had the weak cold resistance. The hybridbetween P. ussuriensis Maxim. and P. bretschneideri Rehd. or P. communis L. had themoderate cold resistance and displayed the central tendency variation, but some individuals displayed super-parent heredity. The resources with the strongest coldresistance could resist minus 45℃.
4. By the evaluation and determination of disease-resistance about winterhardyapple and pear resources, the resources with the strong resistance to apple canker wereM. komarovii, Zha'aishandingzi, Xiaohuanghaitang, Huangtaiping, Xiaosuanguo andDaqiu; the resources with the strong resistance to pear scab were Suanliguozi,Zhenhongxiao and Xifengjingbai, which were very good winterhardy pear germplasmresources. It was also found that the resources which had good cold resistance alsohad the good disease resistance; there was the positive correlation between thecharacters. These results could make a good scientific base for further research,construction of core collections and utilization in breeding about winterhardy appleand pear germplasm resources.
5. By the determination of chromosome ploidy on winterhardy apple and peargermplasm resources, all of the experimental materials belonged to diploid 2n=2x=34, no matter species, fruit or leaf.
6. Based on the results of phenotypic character description, evaluation anddetermination, the database was set up about winterhardy apple and pear germplasmresources.
7. Using the winterhardy apple and pear germplasm resources, RAPD reactionsystem and optimizing reaction procedure were established, 25 primer were selectedand used in 266 apple accessions and 107 pear accessions. The percentage of thepolymorphism bands by RAPD amplifying was 93.47% and 96.20% for apple andpear respectively. By the Nei-Li genetic distance, 180 apple accessions and 87 pearaccessions were analysis and calculated the value of the genetic distance, which was0.0891 to 1.000 in apple (average value 0.00506) and 0.040 to 0.4269 in pear (averagevalue 0.1394). In apple resources, Jinhong4hao and Jinhong7hao had the smallestgenetic distance 0.08911, but M. kansuensis and LingDangGuo had the biggestgenetic distance 1.0000. In pear resources, Suanliguozi and Longliangli had theminimum genetic distance 0.040, however, Heilongjianghonghuagai andYuandongyihao had the maximum genetic distance 0.4269.
8. By the analysis of cluster graph, it was found that Sect. M. baccata, Sect. M.prunifolia and Sect. M. kansuensis had the close genetic relationship which was samewith the classic taxology. But the phenotypic genetic diversity was very rich(abundant) in the Malus resources. Because the phenotypic character between M. baccata and M. prunifolia was close, they had the nearest genetic relationship. It wasverified that M. prunifolia was the hybrid between M. baccata and M. domestica,which was reported before. But there were obvious differences between M.kansuensis or M. komarovii and M. baccata in the phenotypic characters. Combinedwith the analysis of the cluster graph, the present author proposed that M. kansuensisand M. komarovii were derived from the cross of Malus and Crataegus because Malus,Pyrus, Crataegus and Sorbus had the same ancient species. About pear, there wasalso the abundant phenotypic diversity. Because Pingguoli and Kuerlexiangli wereclustered together with P. bretschneideri, and illustrated the near genetic relationshipwith P. bretschneideri, so they were assorted into the system of P. bretschneideri.
9. According to the classic taxology, hybrid progeny in winterhardy appleresources was formed a new series -- hybrid group series in Sect. Malus.
10. According to the results in botanic and biologic investigation, evaluation anddetermination, and the result in RAPD molecular marker, core collections inwinterhardy apple and pear germplasm resources was constructed, which were 46accessions in apple and 29 accessions in pear.
11. Particular RAPD markers were gained from winterhardy apple and peargermplasm resources.
引文
[1]贾敬贤，贾定贤，任庆棉等．中国作物及其野生近缘植物．中国农业出版社，2006
[2]俞德俊．中国果树分类学[M]北京：农业出版社，1979
[3]茹考夫斯基(M育种的世界植物基因资源)．植物育种的遗传学原理[M]．北京：科学出版社，1974
[4]韩振海．落叶果树种质资源学北京：中国农业出版社，1995
[5]孙云尉．中国果树史与果树资源[M]．上海科技出版社，1983
[6]陆秋农，贾定贤，中国果树志．苹果卷，中国农业出版社1999
[7]周恩，印永民，代宝合，等．寒地果树栽培．上海：科技出版社，1982，1-2．
[8]李育农．苹果属植物种质资源研究．中国农业出版社，1989．
[9]蒲富慎，王宇霖．中国果树志(第三卷：梨)．上海：上海科学技术出版社，1963．
[10]藤元文，柴明良，李秀根．梨属植物分类的历史回顾及新进展2004，21(3)252-257．
[11]江用文．国家种质资源圃保存资源名录．中国农业科学技术出版社
[12]李树玲．我国的梨品种资源概况[J]．中国果树，1993(1)：37-39，
[13]王宇霖．半个世纪以来我国梨果产业与科学发展的回顾[J]．果树科学，1999，16(4)：239-245。
[14]顾模．东北中部果树资源的调查北京：科学出版社，1956
[15]顾模．吉林省抗寒果树种质资源的研究与利用．中国农业科学，1984，(3)：37-44
[16]李树玲．我国的梨品种资源概况[J]．中国果树，1993(1)：37-39．
[17]曹玉芬，李树玲，黄礼森，等．我国梨种质资源研究概况及优良种质的综合评价[J]．中国果树2000，(4)：42-44
[18]张冰冰，刘慧涛，宋洪伟等，寒地果树种质资源研究与利用进展．植物遗传资源学报，2006，7(1)：123-128．
[19]张冰冰，刘慧涛，宋洪伟，等．吉林省野生果树种质资源研究综述[J]．吉林农业科学，2005，30(2)：51-54，60
[20]张开春等．分子标记在果树上的应用[J]．果树科学，1999，16(3)：210-218．
[21]Williams F G K, Kubelik A R, Livak K J et al. DNA polymorphisms amplified by arbitrary prim ers are useful as genetic markefs[J]. Nucleic Acids Res, 1990, 18:6531-6535.
[22]Welsh J, Mc Clelland M. Fingerprinting genomes using PCR with arbitrary primers[J]. Nucleic Acids Res, 1990, 18:7213-7218
[23]焦锋，楼程富．RAPD技术应用中的一些问题及对策．
[24]Lanhan PG,Brennan RM, Hackettoc. Mcnicol RJ, RAPD fingerprinting of blackcurrant (Ribes nigrum L) cultivars, Theor. Appl. Genetics, 1995, 90(2): 166-172.
[25]Harada T, Matsukawa K, Sato T, et al.DNA-RAPDs detect genetic variation and paternity in Malus[J]. Euphytica, 1993, 65: 87-91.
[26] 沈向，郭卫东，吴燕民，等．杏43个品种资源的RAPD分类[J]．园艺学报，2000，27(1)：55-56．
[27] Koller B, Lehmann A, McDermott J M, et al. Identification of apple cultivars using RAPD markers [J]. Theor Appl Genet, 1993, 85: 901-904.
[28] Tancred S J, Zeppa A G,Graham G C.The use of PCR-RAPD technique in improving the plant variety fights description of a new Queensland apple(Malus domestica)cultivar[J].Australian Journal of Experimental Agriculture, 1994,34:665-667
[29] Graharm J, et al. Identification of red raspberry cultivars cultivars and an assessment of their relatedness using fingerprints produced by random primers [J]. J Hort Sci, 1994, 69(1): 123-130
[30] 彭建营，束怀瑞，孙仲序．分子标记技术及其在果树种质资源研究上的应用[J]．山东农业大学学报(自然科学版)，2001，32(1)：103-106．
[31] Harada T, Matsukawa K, Sato T, et al. DNA-RAPDs detect genetic variation and paternity in Malus[J]. Euphytica, 1993, 65: 87-91.
[32] 葛颂等．遗传多样性及其检验方法．生物遗传多样性研究的原理与方法．北京：科学出版社，1994，123-140．
[33] 马克平．试论生物多样性的概念[J]．生物多样性，1993，1(1)：20-22。
[34] 沈浩，刘登义．遗传多样性概述[J]．生物学杂志，2001，18(3)：5-9．
[35] 王洪新，胡志昂．植物繁育系统、遗传结构和遗传多样性的保护[J]．生物多样性，1996，4(2)：92-96．
[36] 张开春，吴禄平，李荣旗，等．RAPD技术--检测平邑甜茶遗传一致性的有效方法(简报)[J]．农业生物技术学报，1997，5(2)：201-202．
[37] 达克东，金德敏，伏建民，等．苹果体细胞无性系变异的RAPD评估[J]．核农学报，2001，15(2)：81-85．
[38] 章一华．植物遗传资源研究的一个新领域-核心样品[J]．作物品种资源，1993(2)：1-3．
[39] 高志红．果梅核心种质的构建与分子标记的研究[M]．中国农业大学，2003．
[40] Marsharll P, et al. A simple method to estimate the percentage of hybrity in canola (Brassica napus) Ehybrids. Theor Appl Genet, 1994, 89: 853-858.
[41] Hemmat M, Weeden N F,Manganaris A G, et al. Molecular markers linkage map for apple[J]. J Hered, 1994, 85: 4-11.
[42] Lawson D M, Hemmat M, Weeden N F.The use of mdecular markers to analyze the inheritance of morphological and developmental traits in apple[J].J Amer Soc Hort Sci, 1995,120(3):532-537.
[43] Conner P J, Brown K B, Weeder N F.Randomly amplified polymorphic DNA-based genetic linkage maps of three apple cultivars[J].J Amer Soc Hort Sci, 1997, 122(3): 350-359.
[44] Maliepaard C, Alston F H, Van Arkel G, et al. Aligning male and female linkage maps of apple (Malus pumila Mill) using multi-allelic markers[J].Theor Appl Genet, 1998, 97: 60-73.
[45] Graharm J, et al. Identification of red raspberry cultivars cultivars and an assessment of their relatedness using fingerprints produced by random primers [J]. J Hort Sci, 1994, 69(1): 123-130
[46] 周爱琴，祝军，曲云军，等．应用RAPD技术鉴定苹果砧术[J]．莱阳农学院学报，2000，17(3)：166-169．
[47] 刘孟军．RAPD技术在枣和酸枣种质鉴定上的应用研究[]．韩振海，黄卫东，许雪峰．园艺学论文集[C]．北京：北京农业大学出版社，1995．
[48] 李汝刚．分子标记在苹果鉴定中的应用[J]．生物技术通报，1997(1)：17-21．
[49] 房经贵，章镇，周兰华，等．RAPD标记鉴定果树芽变的可行性分析[J]．果树学报，2001，18(3)：182-185．
[50] 史永忠，郭文武，邓秀新．柑桔RAPD技术体系建立与体细胞杂种鉴定[J]．园艺学报，1998，25(2)：105-110．
[51] Luro F.Laigeretf, Bove Joseph-Marie, et al. DNA amplified fingerprinting, a useful tool for determination of genetic orign and diversity analysis in Citrus[J]. HortScience, 1995, 30(5): 1063-1067.
[52] Harada T, Matsukawa K, Sato T, et al.DNA-RAPDs detect genetic variation and paternity in Malus[J]. Euphytica, 1993, 65: 87-91.
[53] Frankel O. H. and A. H. D. Brown. Current plant genetic resources-a critical appraisal. In: Genetics: new frontiers (vol. Ⅳ). Chopra V. L., Joslu B. C., Sharma R. P.and Bansai H. C. (eds), Oxford & IBH Publ. Co., New Delhi, pp 1984: 1-13.
[54] Van Hintum Th. J. L.. The general methodology for creating a core collection. In: Core collections for today and tomorrow. Johnson R. C. and Hodgkin T.(eds),International Plant Genetic Resources Institute, Rome, Italy, pp1999: 10-17.
[55] Hodgkin T. Core collection and conservation of genetic resouce []. Arora R K, Riley K W. Sesame Bio-diversity in Asia, Conservation, Evaluation and Improment[D]. New Delhi, India, 1993.
[56] Basigalup D H, Barnes D K, Stucker R E .Development of a core collection for perennial Medicago plant Introductions [J]. Crop Science, 1995, 35:1163-1168
[57] Diwan N, Cary R B, Maria S M. A core collection for the US annual Medicago germplasm collection [J]. Crop science, 1994, 34: 279-285.
[58] Diwan N, Mcitosh M S. Bauchan G R. Methods of developing a core collection of annual Medicago species [J]. Theor Appl Genet, 1995, 90: 755-761.
[59] Casler M D. Patterns of variation in a collection of perennial ryegrass accessions [J]. Crop science, 1995, 35:1169-1177
[60] Crossa J, delacy I H, Taba S. The use of multivariate methods in developing a core collection []. Hodgkin T, Brown ADH, Hintum van THL, Morales EAV. Core Collection of Plant Genetic Resources [M]. International Plant Genetic Resource Institute (IPGRI): A Wiley-Sayce Publication, 1995, 77-93
[61] Charmer G,Balfourier F,Ravel C. Genetype × environment interactions in a core collection of French perennial ryegrass populations [J]. Theor Appl Genet, 1993, 86:731-736.
[62] 沈金雄，郭庆元．中国芝麻种质资源的聚类分析[J]．华中农业大学学报，1995，14(6)：532-536．
[63] 李自超，张洪亮，孙传清等．植物遗传资源核心种质研究现状与展望[J]．中国农业大学学报，1999：4(5)：51-62．
[64] Charmet G, Balfourier F,Ravel C., Benis J. B., 1993. Genotype×environment interactions in a core collection of French perennial ryegrass populations. Theor. Appl. Genet., 86:731-736.
[65] Ortiz R., S. Madsen, E. N. Ruiz-Tapia, S. E. Jacobsen, A. Mujica-Sanchez, J. L. Christiansen, O. Stolen. Validating a core collection of Peruvian quinoa germplasm. Genet. Resour. Crop. Evol., 1999, 46: 285-290.
[66] 胡晋．多次聚类构建作物遗传资源核心种质库的方法研究．浙江大学博士学位论文．1999
[67] Abel B. C. and L. M. Pollak, 1991. Rank comparisons of unadapted maize population by testers and per se evaluation. Crop Sci., 31: 650-656.
[68] Bayaa B., W. Erskine & M. Singly, 1997. Screening lentil for resistance to fusarium wilt: methodology and sources of resistance. Euphytica, 98: 69-74.
[69] Boukema I. W., T. J. L. van Hintum and D. Astley, 1997. Creation and composition of the Brassica oleracea core collection. Plant Genetic Resources Newsletter, 111: 29-32.
[70] Brown A H D. Core collection: a practical approach to genetic resources management. Genome, 1989, 31: 818-824.
[71] Miklas P. N., R. Delorme, R. Harman & M. H. Dickson. Using of subsample of the core collection to identify new sources of resistance to white mold in common bean. Crop Sci., 1999, 39: 569-573.
[72] Spagnoletti Zeuli P. L., and C. O. Qualset, 1995. The durum wheat core collection and the plant breeder. In: Core Collections of Plant Genetic Resources. Hodgkin T. et al. (eds), Chichester, UK, John Wiley Sons Publication, pp 213-228.
[73] Frankel O. H. and A. H. D. Brown. Current plant genetic resources-a critical appraisal. In: Genetics: new frontiers (vol.Ⅳ). Chopra V. L., Joslu B. C., Sharma R. P. and Bansal H. C. (eds), Oxford & IBH Publ. Co., New Delhi, pp 1984: 1-13.
[74] Spagnoletti Zeuli P. L. and C. Q. Qualset, 1993. Evaluation of five strategies for obtaining a core subset from a large genetic resource collection of durum wheat. Theor. Appl. Genet., 87: 295-304.
[75] Skroch P. W., J. Nienhuis, S. Beebe, J. Tohme & R Pedraza, 1998. Comparison of mexican common bean (Phaseolus vulgaris L.) core and reserve germplasm collections. Crop Sci., 38:488-496.
[76]Knauft D. A. and Gorbet D. W., 1989. Genetic diversity among peanut cultivars. Crop Sci., 29: 1417-1422．
[77]申时全，曾亚文，李自超等．分子标记及其在云南稻种核心种质中的应用[J]．种子，2001，(6)：8-11．
[78]张秀荣，赵应忠，冯祥运，等．芝麻资源核心收集品的建立[J]．中国油料作物学报，1999，21(3)：27-29
[79]Corley, Holbrook C, William F Anderson. Evaluation of a core collection to identify resistance to late leafspot in peanut [J]. Crop Science, 1995, 35:1700-1702
[80]赵应忠，程勇，张秀荣，等．芝麻资源核心种质与非核心种质的同工酶研究[J]．中国油料作物学报，1998，20(4)：29-34．
[81]彭建营．枣种质资源的分子标记及无核性状的分子标记研究[D]．博士学位论文，1999．
[82]王丙旭．RAPD在梨种质资源亲缘关系和品种鉴定中的应用[M]．吉林农业大学．1998硕士论文．
[83]张广平，李锡香，向长萍等．黄瓜种质核心样本构建方法初探．园艺学报，2006(2)
[84]高志红，章振，韩振海等．中国果梅核心种质的构建与检测．中国农业科学，2005(2)
[85]明军，张启翔，兰彦平等．梅花品种资源核心种质构建．北京林业大学学报，2005(2)
[86]赵丽梅，董英山，刘宝等．中国一年生野生大豆(Glycine soja)核心资源构建．科学通报，2005(10)
[87]李长涛，宋丽丽，孔伟丽等．植物遗传资源核心种质的构建及应用．植物学通报，2005(01)
[88]李天红．桃遗传资源核心种质的研究．中国农学通报．2005(08)
[89]李银霞，高其洁，李天红等．基于果实相关性状的桃品种初级核心种质取样策略研究．果树学报，2006(03)
[90]周少川，李宏湖，黄道强等．水稻核心种质育种．科技导报，2005(11)
[91]李娟，江昌俊等．中国茶树核心种质的初步构建．安徽农业大学学报．2004(03)
[92]邱丽娟，曹永生．常汝镇等．中国大豆核心物质构建．中国农业科学，2003(12)
[93]张洪亮，李自超，曹永生等．表型水平上检验水稻核心种质的参数比较．作物学报，2003(02)
[94]杨生龙，张振海，雍瑞华等．聚类分析在宁夏水稻核心种质研究中的应用．宁夏农林科技，2003(02)
[95]崔艳华，邱丽娟，常汝镇等．植物核心种质研究进展．植物遗传资源学报．2003(03)
[96]董玉琛，曹永生，张学勇等．中国普通小麦初选核心种质的产生．植物遗传资源学报，2003(01)
[97]孙强，林秀云，李明生等．吉林省水稻种资源核心种质构建的研究．吉林农业科学，2006(01)
[98]庄飞云，赵志伟，李锡香等．中国地方胡萝卜品种资源的核心样品构建．园艺学报，2006(01)
[99]刘勇，孙中海，刘德春等．利用分子标记技术选择柚类核心种质资源．果树学报，2006(03)
[100]祝军，周爱琴，王涛，等．苹果RAPD反应体系的建立[J]．果树科学，2000，17(4)：239-243．
[101]曲柏宏、金香兰、陈艳秋、刘洪章、王丕武．梨属种质资源RAPD的分析[J]．园艺学报．2001，28(5)：460-462．
[102]黄礼森，李树玲．我国梨品种染色体数目观察[J]．中国果树，1986，1：12-13
[103]杨槐俊．孢粉学在部分梨属植物分类研究中的应用[J]．果树科学，1985，2(3)：2-9
[104]顾模，钱致斌，孙凤兰等，苹果杂种后代抗寒力遗传规律的研究 园艺学报，1981，8(2)：15-22
[105]钱致斌，顾摸．大苹果与小苹果杂种后代某些遗传倾向的分析．园艺学报，1964，3(2)：139-150
[106]范源洪，蔡青，宿兵，等．DNA提取方法对6种甘蔗亚族植物RAPD的影响[J]．西南农业学报，1999，12(1)：1-7．
[107]张潞生，李传友，贾建航，等．猕猴桃雌雄性别的AFLP鉴别中DNA模板的制备[J]．果树科学，1999，16(3)：171-175．
[108]Oard JH, Dronavalli S. Rapid isolation of rice and maize DNA for analysis by random-primer PCR [J]. Plant molecular Biology Reporter, 1992, 10: 236-241.
[109]卢扬江，郑康乐．提取水稻DNA的一种简易方法[J]．中国水稻科学，1992，6(1)：47-48
[110]王关林，方宏筠，等．植物基因工程原理与技术[D]．北京：科学出版社，1998，370-372
[111]易干军，霍合强，蔡长河等．适于AFLP分析用的荔枝DNA提取方法[[J]．华南农业大学，1999，20(3)：123-124
[112]邹喻苹，汪小金，雷一丁，等．几种濒危植物及其近缘关群总DNA的提取与鉴定[J]．植物学报，1994，36(7)：528-533
[113]Rich U, Schubcrt I. Midiprep method for isolation of DNA from Plants with a high content of polyphenolics [J]. Nucleic Acids Research, 1993, 21(14): 3328
[114]刘平，彭士琪，刘孟军等．分子标记在果树上的应用及前景展望[J]．河北农业大学学报，2002，25(1)：100-105
[115]金勇丰，张耀洲，陈大明，等．桃早熟芽变品种‘大观一号’的RAPD分析及其特异片段的克隆[J]．果树科学，1998，15(2)：103-106
[116]潘新法，孟祥勋，曹广力，等．RAPD在枇杷品种鉴定中的应用[J]．果树学报，2002，19(2)：136-138．
[117]王跃进，Lamikanra O．葡萄RAPD分析影响因子的研究[J]．农业生物技术学报，1997，5(4)：387-391．
[118]文晓鹏，邓秀新．刺梨及其近缘种PCR实验体系的建立与优化[J]．果树学报，2003，20(1)：35-39．
[119]吴少华，张大生，潘东明，等．萘基因组DNA的提取及RAPD条件优化[J]．福建农林大学学报(自然科学版)，2002，31(1)：51-54．
[120]Ballard R, Rajapakse S, Abbott A. DNA markers in rose and their use for cultivar identification and genome mapping [J]. Acta Hort, 1996, 424: 265-268..
[121]李育农．世界苹果属植物的起源演化研究新进展[J]，果树科学，1999(16)：8-19．
[122]王宇霖．落叶果树种类学[M]．北京：农业出版社，1988．
[123]俞德俊．华北的梨(第一版)[M]：科学出版社，1958：17-19．
[124]曲泽洲，孙云蔚．果树种类论．(第一版)[M]：农业出版社，1990：45
[125]曲泽洲，闪崇辉．北京果树志(第一版)[M]：北京出版社，1990：239
[126]林伯年，沈德绪．利用过氧化物酶同工酶分析梨属种质特性及亲缘关系[J]．浙江农业大学学报，1983，9(3)：235-242．
[127]沈允权，李显日，金京国．应用过氧化物酶同工酶谱对苹果梨亲缘关系的研究[J]．延边农学院学报，1992，(2)：91-93．
[128]陕西果树研究所主编．梨主要品种原色谱图．(第一版)[M]：农业出版社，1976：49
[129]原芜洲．西北的梨．(第一版)[M]：陕西科学技术出版社，1979：121-122
[130]中国农业科学院果树研究所．中国果树栽培学[M]．农业出版社，1987
[131]河北农业大学主编．1984果树栽培各论[M]．农业出版社，60．
[132]吴耕民．中国温带果树种类学(第一版)[M]：农业出版社，1984：61
[133]Miklas P. N., R. Delorme, R. Harman & M. H. Dickson. Using of subsample of the core collection to identify new sources of resistance to white mold in common bean. Crop Sci., 1999, 39: 569-573.
[134]Casler M D. Patterns of variation in a collection of perennial ryegrass accessions [J]. Crop science, 1995, 35:1169-1177
[2]俞德俊．中国果树分类学[M]北京：农业出版社，1979
[3]茹考夫斯基(M育种的世界植物基因资源)．植物育种的遗传学原理[M]．北京：科学出版社，1974
[4]韩振海．落叶果树种质资源学北京：中国农业出版社，1995
[5]孙云尉．中国果树史与果树资源[M]．上海科技出版社，1983
[6]陆秋农，贾定贤，中国果树志．苹果卷，中国农业出版社1999
[7]周恩，印永民，代宝合，等．寒地果树栽培．上海：科技出版社，1982，1-2．
[8]李育农．苹果属植物种质资源研究．中国农业出版社，1989．
[9]蒲富慎，王宇霖．中国果树志(第三卷：梨)．上海：上海科学技术出版社，1963．
[10]藤元文，柴明良，李秀根．梨属植物分类的历史回顾及新进展2004，21(3)252-257．
[11]江用文．国家种质资源圃保存资源名录．中国农业科学技术出版社
[12]李树玲．我国的梨品种资源概况[J]．中国果树，1993(1)：37-39，
[13]王宇霖．半个世纪以来我国梨果产业与科学发展的回顾[J]．果树科学，1999，16(4)：239-245。
[14]顾模．东北中部果树资源的调查北京：科学出版社，1956
[15]顾模．吉林省抗寒果树种质资源的研究与利用．中国农业科学，1984，(3)：37-44
[16]李树玲．我国的梨品种资源概况[J]．中国果树，1993(1)：37-39．
[17]曹玉芬，李树玲，黄礼森，等．我国梨种质资源研究概况及优良种质的综合评价[J]．中国果树2000，(4)：42-44
[18]张冰冰，刘慧涛，宋洪伟等，寒地果树种质资源研究与利用进展．植物遗传资源学报，2006，7(1)：123-128．
[19]张冰冰，刘慧涛，宋洪伟，等．吉林省野生果树种质资源研究综述[J]．吉林农业科学，2005，30(2)：51-54，60
[20]张开春等．分子标记在果树上的应用[J]．果树科学，1999，16(3)：210-218．
[21]Williams F G K, Kubelik A R, Livak K J et al. DNA polymorphisms amplified by arbitrary prim ers are useful as genetic markefs[J]. Nucleic Acids Res, 1990, 18:6531-6535.
[22]Welsh J, Mc Clelland M. Fingerprinting genomes using PCR with arbitrary primers[J]. Nucleic Acids Res, 1990, 18:7213-7218
[23]焦锋，楼程富．RAPD技术应用中的一些问题及对策．
[24]Lanhan PG,Brennan RM, Hackettoc. Mcnicol RJ, RAPD fingerprinting of blackcurrant (Ribes nigrum L) cultivars, Theor. Appl. Genetics, 1995, 90(2): 166-172.
[25]Harada T, Matsukawa K, Sato T, et al.DNA-RAPDs detect genetic variation and paternity in Malus[J]. Euphytica, 1993, 65: 87-91.
[26] 沈向，郭卫东，吴燕民，等．杏43个品种资源的RAPD分类[J]．园艺学报，2000，27(1)：55-56．
[27] Koller B, Lehmann A, McDermott J M, et al. Identification of apple cultivars using RAPD markers [J]. Theor Appl Genet, 1993, 85: 901-904.
[28] Tancred S J, Zeppa A G,Graham G C.The use of PCR-RAPD technique in improving the plant variety fights description of a new Queensland apple(Malus domestica)cultivar[J].Australian Journal of Experimental Agriculture, 1994,34:665-667
[29] Graharm J, et al. Identification of red raspberry cultivars cultivars and an assessment of their relatedness using fingerprints produced by random primers [J]. J Hort Sci, 1994, 69(1): 123-130
[30] 彭建营，束怀瑞，孙仲序．分子标记技术及其在果树种质资源研究上的应用[J]．山东农业大学学报(自然科学版)，2001，32(1)：103-106．
[31] Harada T, Matsukawa K, Sato T, et al. DNA-RAPDs detect genetic variation and paternity in Malus[J]. Euphytica, 1993, 65: 87-91.
[32] 葛颂等．遗传多样性及其检验方法．生物遗传多样性研究的原理与方法．北京：科学出版社，1994，123-140．
[33] 马克平．试论生物多样性的概念[J]．生物多样性，1993，1(1)：20-22。
[34] 沈浩，刘登义．遗传多样性概述[J]．生物学杂志，2001，18(3)：5-9．
[35] 王洪新，胡志昂．植物繁育系统、遗传结构和遗传多样性的保护[J]．生物多样性，1996，4(2)：92-96．
[36] 张开春，吴禄平，李荣旗，等．RAPD技术--检测平邑甜茶遗传一致性的有效方法(简报)[J]．农业生物技术学报，1997，5(2)：201-202．
[37] 达克东，金德敏，伏建民，等．苹果体细胞无性系变异的RAPD评估[J]．核农学报，2001，15(2)：81-85．
[38] 章一华．植物遗传资源研究的一个新领域-核心样品[J]．作物品种资源，1993(2)：1-3．
[39] 高志红．果梅核心种质的构建与分子标记的研究[M]．中国农业大学，2003．
[40] Marsharll P, et al. A simple method to estimate the percentage of hybrity in canola (Brassica napus) Ehybrids. Theor Appl Genet, 1994, 89: 853-858.
[41] Hemmat M, Weeden N F,Manganaris A G, et al. Molecular markers linkage map for apple[J]. J Hered, 1994, 85: 4-11.
[42] Lawson D M, Hemmat M, Weeden N F.The use of mdecular markers to analyze the inheritance of morphological and developmental traits in apple[J].J Amer Soc Hort Sci, 1995,120(3):532-537.
[43] Conner P J, Brown K B, Weeder N F.Randomly amplified polymorphic DNA-based genetic linkage maps of three apple cultivars[J].J Amer Soc Hort Sci, 1997, 122(3): 350-359.
[44] Maliepaard C, Alston F H, Van Arkel G, et al. Aligning male and female linkage maps of apple (Malus pumila Mill) using multi-allelic markers[J].Theor Appl Genet, 1998, 97: 60-73.
[45] Graharm J, et al. Identification of red raspberry cultivars cultivars and an assessment of their relatedness using fingerprints produced by random primers [J]. J Hort Sci, 1994, 69(1): 123-130
[46] 周爱琴，祝军，曲云军，等．应用RAPD技术鉴定苹果砧术[J]．莱阳农学院学报，2000，17(3)：166-169．
[47] 刘孟军．RAPD技术在枣和酸枣种质鉴定上的应用研究[]．韩振海，黄卫东，许雪峰．园艺学论文集[C]．北京：北京农业大学出版社，1995．
[48] 李汝刚．分子标记在苹果鉴定中的应用[J]．生物技术通报，1997(1)：17-21．
[49] 房经贵，章镇，周兰华，等．RAPD标记鉴定果树芽变的可行性分析[J]．果树学报，2001，18(3)：182-185．
[50] 史永忠，郭文武，邓秀新．柑桔RAPD技术体系建立与体细胞杂种鉴定[J]．园艺学报，1998，25(2)：105-110．
[51] Luro F.Laigeretf, Bove Joseph-Marie, et al. DNA amplified fingerprinting, a useful tool for determination of genetic orign and diversity analysis in Citrus[J]. HortScience, 1995, 30(5): 1063-1067.
[52] Harada T, Matsukawa K, Sato T, et al.DNA-RAPDs detect genetic variation and paternity in Malus[J]. Euphytica, 1993, 65: 87-91.
[53] Frankel O. H. and A. H. D. Brown. Current plant genetic resources-a critical appraisal. In: Genetics: new frontiers (vol. Ⅳ). Chopra V. L., Joslu B. C., Sharma R. P.and Bansai H. C. (eds), Oxford & IBH Publ. Co., New Delhi, pp 1984: 1-13.
[54] Van Hintum Th. J. L.. The general methodology for creating a core collection. In: Core collections for today and tomorrow. Johnson R. C. and Hodgkin T.(eds),International Plant Genetic Resources Institute, Rome, Italy, pp1999: 10-17.
[55] Hodgkin T. Core collection and conservation of genetic resouce []. Arora R K, Riley K W. Sesame Bio-diversity in Asia, Conservation, Evaluation and Improment[D]. New Delhi, India, 1993.
[56] Basigalup D H, Barnes D K, Stucker R E .Development of a core collection for perennial Medicago plant Introductions [J]. Crop Science, 1995, 35:1163-1168
[57] Diwan N, Cary R B, Maria S M. A core collection for the US annual Medicago germplasm collection [J]. Crop science, 1994, 34: 279-285.
[58] Diwan N, Mcitosh M S. Bauchan G R. Methods of developing a core collection of annual Medicago species [J]. Theor Appl Genet, 1995, 90: 755-761.
[59] Casler M D. Patterns of variation in a collection of perennial ryegrass accessions [J]. Crop science, 1995, 35:1169-1177
[60] Crossa J, delacy I H, Taba S. The use of multivariate methods in developing a core collection []. Hodgkin T, Brown ADH, Hintum van THL, Morales EAV. Core Collection of Plant Genetic Resources [M]. International Plant Genetic Resource Institute (IPGRI): A Wiley-Sayce Publication, 1995, 77-93
[61] Charmer G,Balfourier F,Ravel C. Genetype × environment interactions in a core collection of French perennial ryegrass populations [J]. Theor Appl Genet, 1993, 86:731-736.
[62] 沈金雄，郭庆元．中国芝麻种质资源的聚类分析[J]．华中农业大学学报，1995，14(6)：532-536．
[63] 李自超，张洪亮，孙传清等．植物遗传资源核心种质研究现状与展望[J]．中国农业大学学报，1999：4(5)：51-62．
[64] Charmet G, Balfourier F,Ravel C., Benis J. B., 1993. Genotype×environment interactions in a core collection of French perennial ryegrass populations. Theor. Appl. Genet., 86:731-736.
[65] Ortiz R., S. Madsen, E. N. Ruiz-Tapia, S. E. Jacobsen, A. Mujica-Sanchez, J. L. Christiansen, O. Stolen. Validating a core collection of Peruvian quinoa germplasm. Genet. Resour. Crop. Evol., 1999, 46: 285-290.
[66] 胡晋．多次聚类构建作物遗传资源核心种质库的方法研究．浙江大学博士学位论文．1999
[67] Abel B. C. and L. M. Pollak, 1991. Rank comparisons of unadapted maize population by testers and per se evaluation. Crop Sci., 31: 650-656.
[68] Bayaa B., W. Erskine & M. Singly, 1997. Screening lentil for resistance to fusarium wilt: methodology and sources of resistance. Euphytica, 98: 69-74.
[69] Boukema I. W., T. J. L. van Hintum and D. Astley, 1997. Creation and composition of the Brassica oleracea core collection. Plant Genetic Resources Newsletter, 111: 29-32.
[70] Brown A H D. Core collection: a practical approach to genetic resources management. Genome, 1989, 31: 818-824.
[71] Miklas P. N., R. Delorme, R. Harman & M. H. Dickson. Using of subsample of the core collection to identify new sources of resistance to white mold in common bean. Crop Sci., 1999, 39: 569-573.
[72] Spagnoletti Zeuli P. L., and C. O. Qualset, 1995. The durum wheat core collection and the plant breeder. In: Core Collections of Plant Genetic Resources. Hodgkin T. et al. (eds), Chichester, UK, John Wiley Sons Publication, pp 213-228.
[73] Frankel O. H. and A. H. D. Brown. Current plant genetic resources-a critical appraisal. In: Genetics: new frontiers (vol.Ⅳ). Chopra V. L., Joslu B. C., Sharma R. P. and Bansal H. C. (eds), Oxford & IBH Publ. Co., New Delhi, pp 1984: 1-13.
[74] Spagnoletti Zeuli P. L. and C. Q. Qualset, 1993. Evaluation of five strategies for obtaining a core subset from a large genetic resource collection of durum wheat. Theor. Appl. Genet., 87: 295-304.
[75] Skroch P. W., J. Nienhuis, S. Beebe, J. Tohme & R Pedraza, 1998. Comparison of mexican common bean (Phaseolus vulgaris L.) core and reserve germplasm collections. Crop Sci., 38:488-496.
[76]Knauft D. A. and Gorbet D. W., 1989. Genetic diversity among peanut cultivars. Crop Sci., 29: 1417-1422．
[77]申时全，曾亚文，李自超等．分子标记及其在云南稻种核心种质中的应用[J]．种子，2001，(6)：8-11．
[78]张秀荣，赵应忠，冯祥运，等．芝麻资源核心收集品的建立[J]．中国油料作物学报，1999，21(3)：27-29
[79]Corley, Holbrook C, William F Anderson. Evaluation of a core collection to identify resistance to late leafspot in peanut [J]. Crop Science, 1995, 35:1700-1702
[80]赵应忠，程勇，张秀荣，等．芝麻资源核心种质与非核心种质的同工酶研究[J]．中国油料作物学报，1998，20(4)：29-34．
[81]彭建营．枣种质资源的分子标记及无核性状的分子标记研究[D]．博士学位论文，1999．
[82]王丙旭．RAPD在梨种质资源亲缘关系和品种鉴定中的应用[M]．吉林农业大学．1998硕士论文．
[83]张广平，李锡香，向长萍等．黄瓜种质核心样本构建方法初探．园艺学报，2006(2)
[84]高志红，章振，韩振海等．中国果梅核心种质的构建与检测．中国农业科学，2005(2)
[85]明军，张启翔，兰彦平等．梅花品种资源核心种质构建．北京林业大学学报，2005(2)
[86]赵丽梅，董英山，刘宝等．中国一年生野生大豆(Glycine soja)核心资源构建．科学通报，2005(10)
[87]李长涛，宋丽丽，孔伟丽等．植物遗传资源核心种质的构建及应用．植物学通报，2005(01)
[88]李天红．桃遗传资源核心种质的研究．中国农学通报．2005(08)
[89]李银霞，高其洁，李天红等．基于果实相关性状的桃品种初级核心种质取样策略研究．果树学报，2006(03)
[90]周少川，李宏湖，黄道强等．水稻核心种质育种．科技导报，2005(11)
[91]李娟，江昌俊等．中国茶树核心种质的初步构建．安徽农业大学学报．2004(03)
[92]邱丽娟，曹永生．常汝镇等．中国大豆核心物质构建．中国农业科学，2003(12)
[93]张洪亮，李自超，曹永生等．表型水平上检验水稻核心种质的参数比较．作物学报，2003(02)
[94]杨生龙，张振海，雍瑞华等．聚类分析在宁夏水稻核心种质研究中的应用．宁夏农林科技，2003(02)
[95]崔艳华，邱丽娟，常汝镇等．植物核心种质研究进展．植物遗传资源学报．2003(03)
[96]董玉琛，曹永生，张学勇等．中国普通小麦初选核心种质的产生．植物遗传资源学报，2003(01)
[97]孙强，林秀云，李明生等．吉林省水稻种资源核心种质构建的研究．吉林农业科学，2006(01)
[98]庄飞云，赵志伟，李锡香等．中国地方胡萝卜品种资源的核心样品构建．园艺学报，2006(01)
[99]刘勇，孙中海，刘德春等．利用分子标记技术选择柚类核心种质资源．果树学报，2006(03)
[100]祝军，周爱琴，王涛，等．苹果RAPD反应体系的建立[J]．果树科学，2000，17(4)：239-243．
[101]曲柏宏、金香兰、陈艳秋、刘洪章、王丕武．梨属种质资源RAPD的分析[J]．园艺学报．2001，28(5)：460-462．
[102]黄礼森，李树玲．我国梨品种染色体数目观察[J]．中国果树，1986，1：12-13
[103]杨槐俊．孢粉学在部分梨属植物分类研究中的应用[J]．果树科学，1985，2(3)：2-9
[104]顾模，钱致斌，孙凤兰等，苹果杂种后代抗寒力遗传规律的研究 园艺学报，1981，8(2)：15-22
[105]钱致斌，顾摸．大苹果与小苹果杂种后代某些遗传倾向的分析．园艺学报，1964，3(2)：139-150
[106]范源洪，蔡青，宿兵，等．DNA提取方法对6种甘蔗亚族植物RAPD的影响[J]．西南农业学报，1999，12(1)：1-7．
[107]张潞生，李传友，贾建航，等．猕猴桃雌雄性别的AFLP鉴别中DNA模板的制备[J]．果树科学，1999，16(3)：171-175．
[108]Oard JH, Dronavalli S. Rapid isolation of rice and maize DNA for analysis by random-primer PCR [J]. Plant molecular Biology Reporter, 1992, 10: 236-241.
[109]卢扬江，郑康乐．提取水稻DNA的一种简易方法[J]．中国水稻科学，1992，6(1)：47-48
[110]王关林，方宏筠，等．植物基因工程原理与技术[D]．北京：科学出版社，1998，370-372
[111]易干军，霍合强，蔡长河等．适于AFLP分析用的荔枝DNA提取方法[[J]．华南农业大学，1999，20(3)：123-124
[112]邹喻苹，汪小金，雷一丁，等．几种濒危植物及其近缘关群总DNA的提取与鉴定[J]．植物学报，1994，36(7)：528-533
[113]Rich U, Schubcrt I. Midiprep method for isolation of DNA from Plants with a high content of polyphenolics [J]. Nucleic Acids Research, 1993, 21(14): 3328
[114]刘平，彭士琪，刘孟军等．分子标记在果树上的应用及前景展望[J]．河北农业大学学报，2002，25(1)：100-105
[115]金勇丰，张耀洲，陈大明，等．桃早熟芽变品种‘大观一号’的RAPD分析及其特异片段的克隆[J]．果树科学，1998，15(2)：103-106
[116]潘新法，孟祥勋，曹广力，等．RAPD在枇杷品种鉴定中的应用[J]．果树学报，2002，19(2)：136-138．
[117]王跃进，Lamikanra O．葡萄RAPD分析影响因子的研究[J]．农业生物技术学报，1997，5(4)：387-391．
[118]文晓鹏，邓秀新．刺梨及其近缘种PCR实验体系的建立与优化[J]．果树学报，2003，20(1)：35-39．
[119]吴少华，张大生，潘东明，等．萘基因组DNA的提取及RAPD条件优化[J]．福建农林大学学报(自然科学版)，2002，31(1)：51-54．
[120]Ballard R, Rajapakse S, Abbott A. DNA markers in rose and their use for cultivar identification and genome mapping [J]. Acta Hort, 1996, 424: 265-268..
[121]李育农．世界苹果属植物的起源演化研究新进展[J]，果树科学，1999(16)：8-19．
[122]王宇霖．落叶果树种类学[M]．北京：农业出版社，1988．
[123]俞德俊．华北的梨(第一版)[M]：科学出版社，1958：17-19．
[124]曲泽洲，孙云蔚．果树种类论．(第一版)[M]：农业出版社，1990：45
[125]曲泽洲，闪崇辉．北京果树志(第一版)[M]：北京出版社，1990：239
[126]林伯年，沈德绪．利用过氧化物酶同工酶分析梨属种质特性及亲缘关系[J]．浙江农业大学学报，1983，9(3)：235-242．
[127]沈允权，李显日，金京国．应用过氧化物酶同工酶谱对苹果梨亲缘关系的研究[J]．延边农学院学报，1992，(2)：91-93．
[128]陕西果树研究所主编．梨主要品种原色谱图．(第一版)[M]：农业出版社，1976：49
[129]原芜洲．西北的梨．(第一版)[M]：陕西科学技术出版社，1979：121-122
[130]中国农业科学院果树研究所．中国果树栽培学[M]．农业出版社，1987
[131]河北农业大学主编．1984果树栽培各论[M]．农业出版社，60．
[132]吴耕民．中国温带果树种类学(第一版)[M]：农业出版社，1984：61
[133]Miklas P. N., R. Delorme, R. Harman & M. H. Dickson. Using of subsample of the core collection to identify new sources of resistance to white mold in common bean. Crop Sci., 1999, 39: 569-573.
[134]Casler M D. Patterns of variation in a collection of perennial ryegrass accessions [J]. Crop science, 1995, 35:1169-1177