2011 3, 18(2): 400 406 Journal of Fishery Sciences of China 研究论文 DOI: 10.3724/S.P.J.1118.2011.00400 2n 李晓丽 1,2, 张泽宇 2, 曹淑青 2, 由学策 1., 266003; 2., 116023 2 摘要 : (Undaria pinnatifida)2n,, 2n,, 0.5~1.0 cm, 2n, 10~20, 20 d, 15 20 2n 8.4% 7.4%, 10 1% 20 15 d 2n, 60 d, 2n 100%; 15 30 d 2n, 60 d 38%; 10 45 d 2n, 60 d 12% 2n, 20 10 d 34%, 60 d 100% 2n 3n 2n, [, 2011,18(2): 400 406] : ; ; ; 2n 中图分类号 : S968 文献标识码 : A 文章编号 : 1005 8737 (2011)02 0400 07, [1 10] (Undaria pinnatifida), 2n [11],, 3n 4n,, 3n, [12] 2n 2n, 2n 2n, [8], 3n, 2n 90%, 2n 10%, 2n, 2n 95%, 2n 5% [11],,, 2n 2n,, 2n, 2n, 2n, 3n 收稿日期 : 2010 04 03; 修订日期 : 2010 07 20. 基金项目 : 863 (2006AA10A412). 作者简介 : (1980 ). E-mail:lixiaoli@dlou.edu.cn 通讯作者 :,,. E-mail: zyz@dlou.edu.cn
2 : 2n 401 1 1.1 2n ( І A),, 2n [11], 2n 2n 20, 40 μmol m 2 s 1,, 12 h/d 80 : NaNO 3 10 g/m 3 NaH 2 PO 4 12H 2 O 20 g/m 3 PI 1 L/m 3 3 1 1.2 1.2.1 2n 2n 10 mg( ), 200 ml 120~200 μm, 1 ml (D=12 cm), 20, 50% 2n, 5 1, 2n 1.2.2 2n ( 0.5~1.0 cm),, 20 1 300 ml, 15 1 2n, 2n 50 1.2.3 2n 0.2~0.5 cm( 0.36 cm) 0.5~1.0 cm( 0.78 cm) 1.0~2.0 cm( 1.52 cm) 20 1, 300 ml 20, 2n 1.2.2 1.2.4 2n 2n,,, ( ), 20, ( ) 1 3 1 2n, 2n 2n,, 50 1.2.5 3n (2n ) 1.2.4,, 3 mm 3 1, 20 1.2.6 2n 2n,, Wittman s (1965),,, [1], 10 15 20 ; 1.1, 3 1 1.3 ± ( x SD), SPSS15.0 2 2.1 2n 2n 20 1, 15 d, 2n,, 20 d,,
402 18,, 2~3 ( І B);,, 20, 10 d 200 μm, 1 1mm ( І C), 2, 0.5 cm, 2n 1, 3 2n,, 15 5 d 3.2%, 10 d 5.2%, 20 d 8.4%; 20 5 d 2.6%, 15 d 6.0%, 20 d 7.4%; 10 5 d 2n 0.8%, 10 d 1.0%, 20 d 1.0%, 2.2 2n ( І D),,,,,,,,, 2~3 ( І E), 20, 1, ( І F), 2, ( І G) Tab.1 表 1 温度对裙带菜 2n 配子体孤雌生殖率的影响 Effects of temperature on the parthenogenesis rate of 2n female gametophytes / temperature 5 10 15 20 n=50 ; x SD; % 10 0.8 0.1 ** 1.0 0.2 ** 1.0 0.1 ** 1.0 0.3 ** 15 3.2 0.3 ** 5.2 0.4 ** 7.2 0.3 ** 8.4 0.5 ** 20 2.6 0.2 ** 4.2 0.3 ** 6.0 0.4 ** 7.4 0.2 ** : ** 3 (P<0.01). Note: * * show very significant difference compared among the three groups in one column(p<0.01). 15 μm(12~18 μm), 11 μm(9~12 μm), 2~3, 20 10 d, ( I H), 2n 2n 2n 2 20 15 d 4% 2n, 30 d 46%, 60d 100% 2n,, 10% ; 15 30 d 2n 12%, 60 d 38%, 2n 20 ; 10 45 d 5% 2n, 60 d Tab.2 表 2 温度对裙带菜 2n 配子体诱导率的影响 Effects of temperature on the formation rate of 2n female gametophytes n=50; x ±SD; % / temperature 15 30 45 60 10 0 0 ** 5±1.0 ** 12±1.5 ** 15 0 12±0.8 ** 20±2.0 ** 38±3.0 ** 20 4 46±2.0 ** 72±4.0 ** 100 ** : ** 3 (P<0.01). Note: * * show significant difference among the three groups in one column(p<0.01).
2 : 2n 403 12%, 20 15, 2n 2.3 2n 3,, 0.5~1.0 cm 2n, 15 d 12%, 30 d 74%, 60 d 100%; 1.0~2.0 cm 15 d 2n 6%, 30 d 45%, 60 d 72%; 0.2~0.5 cm 15 d 2n 2%, 30 d 18%, 60d 38%,, 0.5~1.0 cm 2n,, 2n 表 3 裙带菜藻体长度对 2n 配子体诱导率的影响 Tab.3 Effects of sporophytes length on the formation rate of 2n female gametophytes n=20; x ±SD; % /cm sporophytes length 15 30 45 60 0.2 0.5 2±0.3 ** 18±2.0 ** 26±2.4 ** 38±2.0 ** 0.5 1.0 12±2.0 ** 74±4.0 ** 88±4.2 ** 100 1.0 2.0 6±1.5 ** 45±3.5 ** 58±3.0 ** 72±5.0 ** : ** 3 (P<0.01). Note: ** show significant difference among the three groups in one column(p<0.01). 2.4 2n 2n 1,,,,,, 2n, 2n 4~5, 2n 4 10 d, 2n 34%, 15d 84%, 20 d 100%, 表 4 裙带菜 2n 配子体与正常雌配子体成熟率比较 Tab.4 Comparison in maturation rate between 2n and n female gametophyte n=50 ; x ±SD; % group normal female gametophytes 2n 2n female gametophytes 10 15 20 48±2.8 92±3.0 100 34±3.6 84±2.2 100 2n 3n, 2n, 90% 2.5 2n 2, 2 5~6 3n, 3n ( Ι I),, 2n 200 μm,, 3n 5, 10 d 3n 220 μm, 20 d 840 μm, 30 d 1.46 mm( Ι J), 3n 20, 1 1 mm, 2 mm 1, 3.8 mm, 6 mm,, 表 5 裙带菜 3n 幼孢子体与对照组 (2n) 藻体长度比较 Tab.5 Comparison in length between 3n and 2n juvenile sporophytes n=20; x ±SD; mm group 10 20 30 control 0.21±0.03 0.82±0.14 1.40±0.21 3n 3n juvenile sporophytes 0.22±0.05 0.84±0.12 1.46±0.18
404 18 2.6, 2n 60 ( I K), n 30 ( I L) 3 3.1 2n 3n 1,,,, [12], 3n,, 2n, 2n,, 2n [13-16],, 2n,, 2n, 2n, 2n, 3n, 2n,, 2n, 2n, 2n, 2n 2n,, 2n 3.2 2n [7], 2n, 10~20, 2n 2n, 20 2n,, 2n, 20 μmol m 2 s 1 40 μmol m 2 s 1 60 μmol m 2 s 1 2n, 2n,,, 2n, 2n, 2n,, ; 2n, ; 2n 3 0.2~0.5 cm 2n, 1.0~2.0 cm,,, 2n 参考文献 : [1]. [M]. :, 2000: 22 33. [2],,,. [J]., 1998, 28(3): 399 404. [3] Allen S K Jr, Downing S G L. Performance of triploid Pacific oysters, Crassostrea gigas (Thunberg). I. survival, growth, glycogen content and sexual maturation in yearlings [J]. J Exp Mar Biol Ecol, 1986, 102: 197 208.
2 : 2n 405 [4],,. [J]., 21(1): 31 36. [5],. [J]., 2004, 34(3): 53 56. [6] Arai K. Genetic improvement of aquaculture finfish species by chromosome manipulation techniques in Japan [J]. Aquaculture, 2001 (197): 205 228. [7] Tiwary B K, Kirubagaranand R, Ray A K. The biology of triploid fish [J]. Rev Fish Biol Fish, 2004 (14): 391 402. [8] Negri V, Lemmi G. Effect of selection and temperature stress on the production of 2n gametes in Lotus tenuis [J]. Plant Breeding, 1998, 117 (4): 345 349. [9] Ollitrault P, Dambier D, Luro F, et al. Ploidy manipulation for breeding seedless triploid citrus [J].Plant Breed Rev, 2008(20): 323 354. [10] Wendel JF. Genome evolution in polyploids[j]. Plant Mol Biol, 2000(42): 225 249. [11]. [J]., 1999, 6(3): 46 48. [12],,,. 3n 4n [J]., 2007, 31 (3): 350 354. [13],,. [J]., 1979(1): 112 119. [14],. [J]., 1976, 3 (1): 32 38. [15],,,. [J]., 1992, 14 (1): 105 108. [16],,. [J]., 1998, 5: 32 35. Induction of diploid female gametophytes from the vegetative cells of parthenogenetic juvenile sporophytes in Undaria pinnatifida LI Xiaoli 1,2, ZHANG Zeyu 2, CAO Shuqing 2, YOU Xuece 2 1. Key Laboratory of Mariculture, Ministry of Education, Ocean University of China, Qingdao 266003, China; 2. Key Laboratory of Mariculture Biotechnology, Ministry of Agriculture, Dalian Ocean University, Dalian 116023, China Abstract: After being excised into several cells of fragments, the indoor incubated 2n female gametophytes of Undaria pinnatifida matured and produced some juvenile sporophytes without fertilization. Then, the 2n female gametophytes were induced from the vegetative cells of the obtained parthenogenetic 2n juvenile sporophytes, which were removed rhizoids and in a 0.5 1.0 cm length. The parthenogenesis rate of the matured 2n female gametophytes reached 8.4% and 7.4% after 20 days of incubation at 20 and 15, respectively, which were both higher than 1% at 10. The 2n female gametophytes were induced after incubation for 15 days at 20, but for 30 days at 15 and for 45 days at 10. The induction rate of the 2n female gametophytes was 100% at 20, but was 38% at 15 and 12% at 10 after 60 days of incubation, respectively. The maturation rate of the induced 2n female gametophytes, after being excised into fragments, reached 34% after incubation for 10 days and 100% for 20 days at 20, respectively. The 3n juvenile sporophytes were produced by the cross between the induced 2n female gametophyte and the n male gametophyte. In present study a new method of mass production of 2n female gametophytes was founded, which laid a foundation for the scale production of triploid seedlings of Undaria pinnatifida.[journal of Fishery Sciences of China, 2011, 18(2): 400 406] Key words: Undaria pinnatifida; parthnogesis; juvenile sporophtes; 2n female gametophytes Corresponding author: ZHANG Zeyu. E-mail : zyz@dlou.edu.cn
406 中国水产科学 图版 Ι 第 18 卷 裙带菜孤雌生殖藻体 2n 配子体诱导 A. 室内保存培养的 2n 配子体; B. 未受精卵及 2n 孤雌生殖幼孢子体; C. 培养 30 d 的 2n 孤雌生殖幼孢子体; D. 切根诱导的 2n 孤雌生殖幼孢子体; E. 2n 孤雌生殖藻体营养细胞诱导产生 2n 配子体; F. 培养 30 d 后 2n 孤雌生殖藻体表面观; G. 培养 60 d 后形成 2n 配子体藻团的孤雌生殖藻体表面观; H.诱导产生的 2n 配子体成熟; I. 培养 3 d 后的 3n 幼孢子体; J. 培养 30 d 后 的 3n 幼苗;K. 2n 配子体染色体; L. n 配子体染色体. Plate Ι Induction of 2n female gametophytes from the parthenogenetic sporophytes of Undaria pinnatifida A. Indoor cultured 2n female gametophytes; B. Unfertilized eggs and 2n parthenogenesis juvenile sporophytes; C.30-day-old 2n parthenogenetic juvenile sporophytes; D. 2n parthenogenetic juvenile sporophytes with the rhizoids removed; E. Germination of 2n female sporophytes from the vegetative cells of 2n parthenogenetic juvenile sporophytes; F.Surface view of 30-day-old 2n female gametophytes on the frond; G. Surface view of clustered 60-day-old 2n female gametophytes on the frond; H.Maturation of induced 2n female gametophytes; I. 3-day-old 3n juvenile sporophytes; J.30-day-old 3n young seedlings; K.Chromosome of 2n female gameto phyte; L. Chromosome of n female gametophyte.