秋水仙素对金达苜蓿染色体加倍效果的影响
|
摘要
【目的】获得金达苜蓿四倍体的纯合体,为不同倍性苜蓿遗传特性研究及不同倍性间实现基因的流动奠定基础。【方法】以金达苜蓿(Medicago sativa L.cv.Jinda)种子(2n=16)为材料,利用秋水仙素结合组织培养,研究了秋水仙素不同质量浓度(0.02,0.07,0.12mg/mL)、不同处理时间(1,3,7d)对金达苜蓿愈伤组织诱导效果及染色体加倍效果的影响,并从形态学和根尖染色体观察对变异株进行倍性鉴定。【结果】秋水仙素质量浓度和处理时间2个因素对金达苜蓿诱变效果均有一定程度的影响,随着秋水仙素质量浓度的升高、处理时间的延长,愈伤组织的诱导率和分化率均下降,成苗率则规律不明显,其中0.07mg/mL秋水仙素处理3d的诱导效果最好,愈伤组织的诱导率达到60.5%,愈伤组织分化率为33.63%,诱变幼苗成苗率为24.1%。倍性鉴定结果表明,获得的变异植株为四倍体与二倍体的混倍体。【结论】0.07mg/mL秋水仙素处理3d是获得多倍体变异幼苗的最佳条件,获得了四倍体和二倍体的混倍体。
【Objective】This study obtained pure alfalfa tetraploid to provide basis for studying different genetic features of alfalfa and realizing the gene flow between ploidy.【Method】Using colchicine in combination with tissue culturing,the effects of colchicine concentration(0.02,0.07,and 0.12 mg/mL)and treatment time(1,3,and 7 d)on callus induction and chromosome doubling effect of Medicago sativa L.cv.Jinda were studied.The ploidy was also identified from morphology and apical chromosomes.【Result】The concentration of colchicine and treatment time had certain effects on mutation of Medicago sativa L.cv.Jinda.The callus induction rate and regeneration frequency decreased as the increase of concentration of colchicine and treatment time.The treatment of 0.07 mg/mL colchicine for 3 dwas best for polyploidy induction with callus induction rate,regeneration frequency and regeneration rate of 60.5%,33.63% and24.1%,respectively.The ploidy identification showed that the obtained mutant plants were mixoploid.【Conclusion】The treatment with 0.07 mg/mL colchicine for 3 dwas most effective on polyploidy induc-tion,and the mutagenic plant chromosome number was mixoploid including tetraploid and diploid.
【Objective】This study obtained pure alfalfa tetraploid to provide basis for studying different genetic features of alfalfa and realizing the gene flow between ploidy.【Method】Using colchicine in combination with tissue culturing,the effects of colchicine concentration(0.02,0.07,and 0.12 mg/mL)and treatment time(1,3,and 7 d)on callus induction and chromosome doubling effect of Medicago sativa L.cv.Jinda were studied.The ploidy was also identified from morphology and apical chromosomes.【Result】The concentration of colchicine and treatment time had certain effects on mutation of Medicago sativa L.cv.Jinda.The callus induction rate and regeneration frequency decreased as the increase of concentration of colchicine and treatment time.The treatment of 0.07 mg/mL colchicine for 3 dwas best for polyploidy induction with callus induction rate,regeneration frequency and regeneration rate of 60.5%,33.63% and24.1%,respectively.The ploidy identification showed that the obtained mutant plants were mixoploid.【Conclusion】The treatment with 0.07 mg/mL colchicine for 3 dwas most effective on polyploidy induc-tion,and the mutagenic plant chromosome number was mixoploid including tetraploid and diploid.
引文
[1]刘卢生,王友国,玉永雄,等.紫花苜蓿倍性研究进展[J].草业科学,2006,23(2):9-14.Liu L S,Wang Y G,Yu Y X,et al.Advances in alfalfa ploidy studies[J].Pratacultural Science,2006,23(2):9-14.
[2]云岚,云锦凤,李俊琴,等.新麦草愈伤组织多倍体诱导与倍性鉴定[J].草业学报,2010,19(6):126-131.Yun L,Yun J F,Li J Q,et al.Callus polyploidy induction and identification of Russian wild ryegrass[J].Acta Pratacultural Sinica,2010,19(6):126-131.
[3]Yang Z,Shen Z,Tetreault H,et al.Production of autopolyploid lowland switchgrass lines through in vitro chromosome doubling[J].Bioenergy Research,2014,7(1):232-242.
[4]Dong Y,Zhao W,Li X,et al.Androgenesis,gynogenesis,and parthenogenesis haploids in cucurbit species[J].Plant Cell Reports,2016,35(10):1991-2019.
[5]Gomes S S L,Saldanha C W,Neves C S,et al.Karyotype,genome size,and in vitro chromosome doubling of Pfaffia glomerata(Spreng.)Pedersen[J].Plant Cell,Tissue and Organ Culture,2014,118(1):45-56.
[6]Würschum T,Tucker M R,Reif J C,et al.Improved efficiency of doubled haploid generation in hexaploid triticale by in vitro chromosome doubling[J].BMC Plant Biology,2012,12(1):109.
[7]王璐.秋水仙素对直立型扁蓿豆诱变效应的研究[D].呼和浩特:内蒙古农业大学,2014.Wang L.Study on inductive effects of Medicago ruthenic(L.)sojak.cv.‘Zhilixing’by colchicine induction[D].Hohhot:Inner Mongolia Agricultural University,2014.
[8]孔舒颖.扁蓿豆新品系的多倍体诱导研究[D].呼和浩特:内蒙古农业大学,2010.Kong S Y.Studies on polyploid induction of Melilotoides ruthenica strains[D].Hohhot:Inner Mongolia Agricultural University,2010.
[9]李悦,师尚礼.秋水仙素处理清水苜蓿胚根对染色体倍性与叶表皮细胞的诱变效应[J].草业学报,2016,25(2):141-149.Li Y,Shi S L.The mutagenic effects of colchicine treatment on chromosome ploidy and the leaf epidermal cells of radicles of Medicago sativa cv.Qingshui[J].Acta Pratacultural Sinica,2016,25(2):141-149.
[10]赵海明,谢楠,李源,等.秋水仙素对染色体加倍研究及在牧草育种上的应用[C]//中国草学会.中国草学会青年工作委员会学术研讨会论文集.[出版地不详]:中国草学会,2007.Zhao H M,Xie N,Li Y,et al.Colchicine for chromosome doubling research and application in forage breeding[C]//Chinese Society of Grassland.The Chinese society of grass academic symposium on youth work committee.[S.l.]:Chinese Society of Grassland,2007.
[11]宋策,陈典,梁誉,等.秋水仙素离体诱导分蘖洋葱茎尖多倍体的研究[J].吉林农业大学学报,2016,38(6):675-680.Song C,Chen D,Liang Y,et al.In vitroinduction of polyploid by colchicine in shoot tips of tillered-onion[J].Journal of Jilin Agricultural University,2016,38(6):675-680.
[12]Saunders J W,Bingham E T.Production of alfalfa plants from tissue culture[J].Crop Science,1972(12):804-808.
[13]李鹏飞.西瓜多倍体诱导及其分子生物学基础[D].河北保定:河北农业大学,2013.Li P F.Studies on the polyploid watermelon induction and the foundations of molecular biology[D].Baoding,Hebei:Hebei Agricultural University,2013.
[14]宁强.枣愈伤组织的诱导及多倍体诱变研究[D].河北保定:河北农业大学,2006.Ning Q.Reseacrh on callus induction and poly ploid mutagenesis in Ziziphus jujube Mill.[D].Baoding,Hebei:Hebei Agricultural University,2006.
[15]钟声,黄梅芬,段新慧.野生二倍体鸭茅染色体加倍及杂交利用研究[J].热带农业工程,2009,33(3):40-43.Zhong S,Huang M F,Duan X H.Chromosome doubling and crossing of wild diploid Dactylis glomerata[J].Tropical Agricultural Engineering,2009,33(3):40-43.
[16]胡瑞阳,段红静,林华忠,等.杉木多倍体变异苗诱导及倍性鉴定[J].核农学报,2016,30(8):1491-1497.Hu R Y,Duan H J,Li H Z,et al.Polyploid mutant seedlings induction and ploidy identification of Chinese fir[J].Journal of Nuclear Agricultural Sciences,2016,30(8):1491-1497.
[17]刘洋,杨利平.朝鲜百合离体多倍体诱导[J].河北农业大学学报,2015,38(3):30-33.Liu Y,Yang L P.Polyploidy induction of Lilium amabile in vitro[J].Journal of Agricultural University of Hebei,2015,38(3):30-33.
[18]Zahedi A A,Hosseini B,Fattahi M,et al.Overproduction of valuable methoxylated flavones in induced tetraploid plants of Dracocephalum kotschyi[J].Botanical Studies,2014,55(1):1-10.
[19]郝峰,刘翠玲,郝水源.蒙古冰草与航道冰草杂交F1代染色体加倍植株过氧化物同工酶分析[J].河套大学学报,2010,30(4):1-3.Hao F,Liu C L,Hao S Y.An analysis of reciprocal hybrids F1between agropyron mongolicun and A.Cristatumcv.Fairway and its chromosome doubling plant isozyme[J].Jurnal of Hetao University,2010,30(4):1-3.
[20]刘辉.马铃薯花药培养及双单倍体植株的鉴定[D].黑龙江哈尔滨:东北农业大学,2009.Liu H.Anthetr culture and indentification of regenertated plants in potato[D].Harbin,Heilongjiang:The Northeast Agricultural University,2009.
[21]秦素平,陈于和,林小虎,等.秋水仙素处理对黑麦根尖细胞有丝分裂的影响[J].麦类作物学报,2006,26(4):142-144.Qin S P,Chen Y H,Lin X H,et al.Effect of colchicine on mitosis in root tip cell of rye(Secale cereal)[J].Journal of Triticeae Crops,2006,26(4):142-144.
[22]刘倩.6种苜蓿属植物多倍体诱导及倍性鉴定研究[D].江苏扬州:扬州大学,2014.Liu Q.Polyploidy induction and ploidy identification of 6Medicago spp.Germplasms[D].Yangzhou,Jiangsu:Yangzhou University,2014.
[23]程金水.园林植物遗传育种学[M].北京:中国林业出版社,2000.Cheng J S.Garden plant genetic breeding[M].Beijing:China Forestry Press,2000.
[24]房永雨,谢锐,于肖夏,等.秋水仙素处理高丹草杂种F1种子诱变效果分析[J].中国草地学报,2013,35(6):40-45.Fang Y Y,Xie R,Yu X X,et al.Analysis of mutagenic effect of colchicines on seeds of sorghum-sudangrass hybrid F1[J].Chinese Journal of Grassland,2013,35(6):40-45.
[25]Mori S,Yamane T,Yahata M,et al.Chromosome doubling in Limonium bellidifolium(Gouan)Dumort.by colchicine treatment of seeds[J].The Horticulture Journal,2016,doi:10.2503/hortj.MI-117.
[26]李洁,石凤翎,卞晓燕,等.秋水仙素诱导直立型扁蓿豆多倍体的研究[C]//中国草学会牧草育种委员会.中国草学会牧草育种委员会第七届代表大会论文集.[出版地不详]:中国草学会牧草育种委员会,2009.Li J,Shi F L,Bian X Y,et al.Polyploidy induction of Melilotoides ruthenicus(L.)Sojak cv.Zhilixing by colchicines[C]//China Grass Society Pasture Breeding Committee.The Chinese society of grass forage breeding committee proceedings of the seventh congress.[S.l.]:China Grass Society Pasture Breeding Committee,2009.
[27]张颖.羊草与灰色赖草杂种F1再生体系建立及染色体加倍的研究[D].呼和浩特:内蒙古农业大学,2007.Zhang Y.Studies on system of plant regeneration and chromosome doubling of the hybrid between Leymus chinensis and Leymus cinereus[D].Hohhot:Inner Mongolia Agricultural University,2007.
[2]云岚,云锦凤,李俊琴,等.新麦草愈伤组织多倍体诱导与倍性鉴定[J].草业学报,2010,19(6):126-131.Yun L,Yun J F,Li J Q,et al.Callus polyploidy induction and identification of Russian wild ryegrass[J].Acta Pratacultural Sinica,2010,19(6):126-131.
[3]Yang Z,Shen Z,Tetreault H,et al.Production of autopolyploid lowland switchgrass lines through in vitro chromosome doubling[J].Bioenergy Research,2014,7(1):232-242.
[4]Dong Y,Zhao W,Li X,et al.Androgenesis,gynogenesis,and parthenogenesis haploids in cucurbit species[J].Plant Cell Reports,2016,35(10):1991-2019.
[5]Gomes S S L,Saldanha C W,Neves C S,et al.Karyotype,genome size,and in vitro chromosome doubling of Pfaffia glomerata(Spreng.)Pedersen[J].Plant Cell,Tissue and Organ Culture,2014,118(1):45-56.
[6]Würschum T,Tucker M R,Reif J C,et al.Improved efficiency of doubled haploid generation in hexaploid triticale by in vitro chromosome doubling[J].BMC Plant Biology,2012,12(1):109.
[7]王璐.秋水仙素对直立型扁蓿豆诱变效应的研究[D].呼和浩特:内蒙古农业大学,2014.Wang L.Study on inductive effects of Medicago ruthenic(L.)sojak.cv.‘Zhilixing’by colchicine induction[D].Hohhot:Inner Mongolia Agricultural University,2014.
[8]孔舒颖.扁蓿豆新品系的多倍体诱导研究[D].呼和浩特:内蒙古农业大学,2010.Kong S Y.Studies on polyploid induction of Melilotoides ruthenica strains[D].Hohhot:Inner Mongolia Agricultural University,2010.
[9]李悦,师尚礼.秋水仙素处理清水苜蓿胚根对染色体倍性与叶表皮细胞的诱变效应[J].草业学报,2016,25(2):141-149.Li Y,Shi S L.The mutagenic effects of colchicine treatment on chromosome ploidy and the leaf epidermal cells of radicles of Medicago sativa cv.Qingshui[J].Acta Pratacultural Sinica,2016,25(2):141-149.
[10]赵海明,谢楠,李源,等.秋水仙素对染色体加倍研究及在牧草育种上的应用[C]//中国草学会.中国草学会青年工作委员会学术研讨会论文集.[出版地不详]:中国草学会,2007.Zhao H M,Xie N,Li Y,et al.Colchicine for chromosome doubling research and application in forage breeding[C]//Chinese Society of Grassland.The Chinese society of grass academic symposium on youth work committee.[S.l.]:Chinese Society of Grassland,2007.
[11]宋策,陈典,梁誉,等.秋水仙素离体诱导分蘖洋葱茎尖多倍体的研究[J].吉林农业大学学报,2016,38(6):675-680.Song C,Chen D,Liang Y,et al.In vitroinduction of polyploid by colchicine in shoot tips of tillered-onion[J].Journal of Jilin Agricultural University,2016,38(6):675-680.
[12]Saunders J W,Bingham E T.Production of alfalfa plants from tissue culture[J].Crop Science,1972(12):804-808.
[13]李鹏飞.西瓜多倍体诱导及其分子生物学基础[D].河北保定:河北农业大学,2013.Li P F.Studies on the polyploid watermelon induction and the foundations of molecular biology[D].Baoding,Hebei:Hebei Agricultural University,2013.
[14]宁强.枣愈伤组织的诱导及多倍体诱变研究[D].河北保定:河北农业大学,2006.Ning Q.Reseacrh on callus induction and poly ploid mutagenesis in Ziziphus jujube Mill.[D].Baoding,Hebei:Hebei Agricultural University,2006.
[15]钟声,黄梅芬,段新慧.野生二倍体鸭茅染色体加倍及杂交利用研究[J].热带农业工程,2009,33(3):40-43.Zhong S,Huang M F,Duan X H.Chromosome doubling and crossing of wild diploid Dactylis glomerata[J].Tropical Agricultural Engineering,2009,33(3):40-43.
[16]胡瑞阳,段红静,林华忠,等.杉木多倍体变异苗诱导及倍性鉴定[J].核农学报,2016,30(8):1491-1497.Hu R Y,Duan H J,Li H Z,et al.Polyploid mutant seedlings induction and ploidy identification of Chinese fir[J].Journal of Nuclear Agricultural Sciences,2016,30(8):1491-1497.
[17]刘洋,杨利平.朝鲜百合离体多倍体诱导[J].河北农业大学学报,2015,38(3):30-33.Liu Y,Yang L P.Polyploidy induction of Lilium amabile in vitro[J].Journal of Agricultural University of Hebei,2015,38(3):30-33.
[18]Zahedi A A,Hosseini B,Fattahi M,et al.Overproduction of valuable methoxylated flavones in induced tetraploid plants of Dracocephalum kotschyi[J].Botanical Studies,2014,55(1):1-10.
[19]郝峰,刘翠玲,郝水源.蒙古冰草与航道冰草杂交F1代染色体加倍植株过氧化物同工酶分析[J].河套大学学报,2010,30(4):1-3.Hao F,Liu C L,Hao S Y.An analysis of reciprocal hybrids F1between agropyron mongolicun and A.Cristatumcv.Fairway and its chromosome doubling plant isozyme[J].Jurnal of Hetao University,2010,30(4):1-3.
[20]刘辉.马铃薯花药培养及双单倍体植株的鉴定[D].黑龙江哈尔滨:东北农业大学,2009.Liu H.Anthetr culture and indentification of regenertated plants in potato[D].Harbin,Heilongjiang:The Northeast Agricultural University,2009.
[21]秦素平,陈于和,林小虎,等.秋水仙素处理对黑麦根尖细胞有丝分裂的影响[J].麦类作物学报,2006,26(4):142-144.Qin S P,Chen Y H,Lin X H,et al.Effect of colchicine on mitosis in root tip cell of rye(Secale cereal)[J].Journal of Triticeae Crops,2006,26(4):142-144.
[22]刘倩.6种苜蓿属植物多倍体诱导及倍性鉴定研究[D].江苏扬州:扬州大学,2014.Liu Q.Polyploidy induction and ploidy identification of 6Medicago spp.Germplasms[D].Yangzhou,Jiangsu:Yangzhou University,2014.
[23]程金水.园林植物遗传育种学[M].北京:中国林业出版社,2000.Cheng J S.Garden plant genetic breeding[M].Beijing:China Forestry Press,2000.
[24]房永雨,谢锐,于肖夏,等.秋水仙素处理高丹草杂种F1种子诱变效果分析[J].中国草地学报,2013,35(6):40-45.Fang Y Y,Xie R,Yu X X,et al.Analysis of mutagenic effect of colchicines on seeds of sorghum-sudangrass hybrid F1[J].Chinese Journal of Grassland,2013,35(6):40-45.
[25]Mori S,Yamane T,Yahata M,et al.Chromosome doubling in Limonium bellidifolium(Gouan)Dumort.by colchicine treatment of seeds[J].The Horticulture Journal,2016,doi:10.2503/hortj.MI-117.
[26]李洁,石凤翎,卞晓燕,等.秋水仙素诱导直立型扁蓿豆多倍体的研究[C]//中国草学会牧草育种委员会.中国草学会牧草育种委员会第七届代表大会论文集.[出版地不详]:中国草学会牧草育种委员会,2009.Li J,Shi F L,Bian X Y,et al.Polyploidy induction of Melilotoides ruthenicus(L.)Sojak cv.Zhilixing by colchicines[C]//China Grass Society Pasture Breeding Committee.The Chinese society of grass forage breeding committee proceedings of the seventh congress.[S.l.]:China Grass Society Pasture Breeding Committee,2009.
[27]张颖.羊草与灰色赖草杂种F1再生体系建立及染色体加倍的研究[D].呼和浩特:内蒙古农业大学,2007.Zhang Y.Studies on system of plant regeneration and chromosome doubling of the hybrid between Leymus chinensis and Leymus cinereus[D].Hohhot:Inner Mongolia Agricultural University,2007.