2007, 34 (1) : 47-52 Acta Horticulturae Sinica SS 1, 23, 13, 2, 1, 3 ( 1,, 310029; 2, 650205; 3, 430209) : SS ( Simp le sequence repeat) 29( ), 6 SS (BGA35 KU10 BGT23b NH004a NH011b NH015a) NH004a, 3, 6 29() 4, 2,,,,, : ; SS; ; : S 66112: A : 05132353X (2007) 0120047206 S im ple Sequence epea t Ana lysis on Genetic A ssessm en t of Ch inese ed Sk inned Sand Pear Cultivars ZHANG Dong 1, SHU Qun 23, TENG Yuan2wen 13, Q IU M ing2hua 2, BAO Lu 1, and HU Hong2ju 3 ( 1 D epartm ent of Horticulture, Zhejiang U niversity, the S tate A gricultural M inistry Laboratory of Horticultural P lant Grow th, D evelopm ent & B iotechnology, Hangzhou 310029, China; Kunm ing 650205, China; : 2006-04 - 01; : 2006-11 - 21 : (30370988) ; (2003NG13) 2 Institute of Horticulture, Yunnan A cadem y of A gricultural Sciences, 3 Institute of F riut and Tea, Hubei A cadem y of A gricultural Sciences, W uhan 430209, China) Abstract: A total of 29 sand pear germp lasm (mainly red skinned) native to southwest of China were subjected to simp le sequence repeat ( SS ) analysis. Six pairs of SS p rimers (B GA35, KU10, B GT23b, NH004a, NH011b and NH015a) could generate a large number of alleles. The p rimer NH004a, which p ro2 duced the highest allele numbers and the most effective alleles, and high heterozygosity and Shannon informa2 tion index, showed the best identification power. U sing six pairs of SS p rimers, all accessions or types could be distinguished excep t for three pairs of cultivars, which m ight belong to the synonymy cases or bud mutants. Twenty nine pear accessions could be divided into four major group s obviously based on the UPGMA cluster a2 nalysis. Group consisted of cultivars all native to Yunnan p rovince, including two green and nine red skinned sand pears, inferring a near genetic relationship between the red and green skinned pears. In group s and, the red skinned pears from Huili County of Sichuan Province and from Yunnan Province m ingled to2 gether, which m ight suggest cultivar movement from Yuannan to Sichuan. In group, not only were Xiangsu2 li and Zaiyangli from Huili County of Sichuan Province, M idu Xiangsuli and Changshui Huobali native to Yun2 nan Province not closely clustered, but also were far from other group s. Huobali pear types distributed in Yun2 nan and Sichuan p rovinces m ight have different origins. The results above indicates large genetic diversity of germp lasm resources of red skinned sand pear cultivars in China. Key words: Sand pear; SS; Cultivar identification; Genetic relationship 3 Author for correspondence ( E2mail: ywteng@ zju1edu1cn; shuqun@public1km1vn1cn)
48 34 ( Kajiura, 1978), (, 1993, 1997) (, 2004) (1997) 14,,, SS ( Sim2 p le sequence repeat) ( Powell et al., 1996 ), ( Kimura et al., 2002; Yamamoto et al., 2002) SS,, 1 111 (H IFT) (CPG) ( YNH I) ( ZZF I) No. Cultivars or lines ( 1) 1 Table 1 Accession s used in th is study O rigin ed skin 1 ()Mantianhong ( Kousui Huobali) Zhengzhou, Henan Yes YNH I 2 ()Meirensu ( Kousui Huobali) Zhengzhou, Henan Yes YNH I 3 1 Xionggu Huobali 1 L ijiang, Yunnan Yes H IFT 4 2 Huili 2 Huili, Sichuan Yes YNH I 5 Suomei L ijiang, Yunnan No ZZF I 6 Baozhuli Chenggong, Yunnan No CPG 7 1 Huili 1 Huili, Sichuan Yes YNH I 8 Changshui Huobali L ijiang, Yunnan Yes H IFT 9 L ijiangm ianli L ijiang, Yunnan Yes H IFT 10 2 Xionggu Huobali 2 L ijiang, Yunnan Yes H IFT 11 Yanshan Hongli Yanshan, Yunnan Yes YNH I 12 2 Yunhongli 2 W enshan, Yunnan Yes H IFT 13 W enshan Hongli W enshan, Yunnan Yes H IFT 14 L ijiang Baili L ijiang, Yunnan No ZZF I 15 1 Yunhongli 1 Yanshan, Yunnan Yes ZZF I 16 M idu Huobali M idu, Yunnan Yes H IFT 17 M idu Xiangsuli M idu, Yunnan Yes H IFT 18 Piaoxing Huobali Huili, Sichuan Yes YNH I 19 M idu Xiaohongli M idu, Yunnan Yes H IFT 20 Laoyali Huili, Sichuan Yes YNH I 21 W enshan Hongxueli W enshan, Yunnan Yes ZZF I 22 Caim ian Yali Huili, Sichuan Yes YNH I 23 Xiangsuli Huili, Sichuan Yes YNH I 24 Dali Huobali Dali, Yunnan Yes H IFT 25 Zaiyangli Huili, Sichuan Yes YNH I 26 Dali Yanzhili Dali, Yunnan Yes H IFT 27 Changba Huobali Huili, Sichuan Yes YNH I 28 Huobali W uding, Yunnan Yes H IFT 29 W eishan Hongxueli W eishan, Yunnan Yes YNH I Sampling source
1 : SS 49 112 11211 DNA SDS DNA,, (Beckman DU800, USA ) DNA 10 ng/ L BGA35 NH011b BGT23b NH0 15a KU10 NH004a ( 2) Yamamoto (2002) SS ( Kimura et al., 2002), Invitrogen TM 11212 PC 20 L, 110 L (10 ng/ L) DNA 210 L 10 buffer (MgCl 2 115 mmol/l ) 210 L 215 mmol/l dntps 110 L p rimer (10 pmoles) Primers 2 SS Table 2 SS pr im er sequences and a llele size ranges Sequences BGA35 F AGAGGGAGAAAGGCGATT 121 160 GCTTCATCACCGTCTGCT KU10 F AGTATGTGACCACCCCGATGTT 222 274 AGAGTCGGTTGGGAAATGATTG BGT23b F CACATTCAAAGATTAAGAT 192 215 ACTCAGCCTTTTTTTCCCAC Allele size ranges(bp) NH004a F AGGATGGGACGAGTT TAGAG 78 126 NH011b F CCACATCTCTCAACCTACCA TGGTTCACATAGAGAGAGAGAGAGA 160 240 TTTGCCGTTGGACCGAGC NH015a F TTGTGCCCTTTTTCCTACC 105 180 CTTTGATGTTACCCCTTGCTG 012 L Taq polymerase ( Takara B iotechnology Company, Japan) 1318 L ddh 2 O, NH011b NH015a NH004a 3 SS: 94 2 m in, 94 1 m in, 55 1 m in 72 2 m in, 35 BGA35 BGT23b KU10 : 94 2 m in, 94 1 m in, 60 1 m in 72 2 m in, 10, 015, 94 2 m in, 94 5 m in, 2 m in, 35 ( Kimura et al., 2002) 55 1 m in 72, PC 110%, 100 200 bp,, ( 1 mg, 1 mg, 015 mol/l EDTA, ), 94 5 m in (Vos et al., 1995), 4 11213 5 L 6% 110 h (75 W ), Bassam (1991) ( 1 0), Nei L i (1979) D ice,, NTSYS2pc, (UPGMA ) SS( ), POPGENE version 1131 SS( n o ) SS ( n e ) (H o ) (H e ) ( I) 2 6 SS( ), NH004a, KU10 BGA35a 7( 3) NH004a, 1 2( 1) 3, 6, H e 012861 018699, 016788 H o 012069 110000, 016839 I 016918 211674, 115005 ( 3) Locus 3 SS Table 3 The character iza tion of polym orph ic SS loci of red sk inned sand pears n o n e H o H e I BGT23b 9 113912 012069 012861 016918 NH011b 8 219561 016897 016733 113875 BGA35a 7 313911 019655 017175 113962 KU10 7 211674 018621 018699 211674 NH015a 11 315940 013793 017344 115431 NH004a 12 414973 110000 017913 118169 Mean 9 219995 016839 016788 115005
50 34 1NH011b 610% A: ; B: 1 F ig. 1 610% PAGE gel electrophoresis of am plif ica tion products w ith NH011b pr im er A rrow A: The specific band of Changba Huobali; A rrow B: The specific band of Zaiyangli. Numbers refer to the cultivars listed in Table 1. D ice, UPGMA ( 2) 4 11,, 3 1 ( ) 0193, 2, 1,, 0192 SS, 1 6 SS, 2 1 2,, 1 2 0189, 0176, 2 UPGM A29 F ig. 2 The phylogenetic tree of 29 sand pear cultivars ( ma in ly red sk inned) ba sed on UPGM A cluster ana lysis 0185, 2 1,,, 0186,
1 : SS 51, 8, ( 2) 1, 013200 ( 2 ) 018889 ( 1 2 ) ( 4) 4 8 D ice Table 4 D iceπs sim ilarity coeff icien ts for 8 Huoba li pear cultivars from d ifferent places No. 27 8 24 28 18 17 3 8 014348 24 015217 015217 28 015217 015217 110000 18 014615 014800 016400 016400 17 014167 014348 016087 016087 0184623 3 015600 013333 015000 015000 015185 014800 10 015385 013200 014800 014800 015000 014615 018889 3 SS ( Zhebentyayeta et al., 2003) SS9, Kimura (2002) 1418, (H o ) 0168, Kimura (2002) 0163, 6 SS,, NH004a Kimura (2002) 9SS660, 3,, 1 2 ( 2) 1 1984, (, 2003) SS, SS 2,,,,,,, 1 2 ( 4), ( 4) ( 2),,, (, ), ( 2), (, 1993), 0193,, 1, ( 2),,,
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