广藿香毛状根多倍体诱导及其植株再生

为了提高药用植物广藿香的次生物质广藿香醇含量,采用秋水仙素人工诱导染色体加倍技术,进行了广藿香毛状根多倍体诱导及其植株再生、倍性鉴定和挥发油组分广藿香醇含量的测定。结果表明,广藿香毛状根多倍体诱导的最佳条件为0.05%秋水仙素处理36 h,其多倍体诱导率可达40%以上;经秋水仙素加倍的广藿香毛状根在MS+6-BA 0.2 mg/L+NAA 0.1 mg/L培养基中培养60 d后可获得毛状根多倍体再生植株。与对照(二倍体植株)相比,广藿香毛状根多倍体再生植株根系更发达、茎更粗、节间变短、叶片的长度、宽度和厚度均较二倍体明显增大。根尖细胞染色体压片观察证实,所获得的广藿香毛状根多倍体再生植株为四倍体,其根尖细胞染色体数约为128;同时,其叶片的气孔保卫细胞体积及其叶绿体数目均约为对照的两倍;但其气孔密度则随着倍性增加而下降,二倍体植株叶片的气孔密度约为四倍体植株叶片的1.67倍。GC-MS测定结果表明,广藿香毛状根多倍体再生植株的广藿香挥发油组分广藿香醇的含量为4.25 mg/g干重,约为二倍体植株的2.30倍。该结果证实毛状根多倍体化可提高药用植物广藿香的广藿香醇含量。     

大花蕙兰多倍体的高效诱导

以大花蕙兰‘红瀑布’无菌苗丛芽为材料、秋水仙素为诱变剂,采用不同的处理浓度、时间诱导大花蕙兰体细胞加倍。通过形态学和细胞学观察、统计等方法对其进行倍性鉴定。结果表明：秋水仙素浓度0.05%,处理时间24 h的条件下,诱导率高达28.2%;多倍体苗外部形态、叶绿体数目、气孔数目和大小与二倍体差异大,加倍后的细胞核明显变大,染色体倍数增加。     

滇北球花报春多倍体诱导及核型分析

以滇北球花报春(Primula denticulata ssp.sinodenticulata)为供试材料,在离体条件下,采用秋水仙素对其丛生芽进行诱导,比较不同浓度、不同处理时间秋水仙素诱导多倍体的效果.结果表明:以0.6%的秋水仙素处理72 h诱导效果最佳,诱导率达54%.经形态学观察发现,变异材料叶色变深,叶片质感变厚;气孔面积增大,单位面积气孔数目减少;染色体计数及核型分析显示,滇北球花报春的二倍体核型为2n=2x=4m+16sm+2st,四倍体核型为2n=4x=8m+32sm+4st,均属3A核型,并成功获得了滇北球花报春的四倍体植株.     

十六倍体三叶半夏品种特性的分析

利用三叶半夏悬浮细胞为材料,通过秋水仙素诱导后获得稳定的半夏多倍体株系。通过染色体鉴定、生理分析以及药典规定项目的测定,综合考察半夏多倍体株系的品种特性,评价其作为新品种的潜力。结果表明:优良半夏株系为八倍体三叶半夏,加倍后的半夏为染色体加倍的十六倍体半夏。与八倍体半夏相比,十六倍体半夏在形态学方面表现出植株矮壮,叶片变圆变厚,块茎增大的特点,细胞学方面表现出气孔密度降低,保卫细胞增大,叶绿体个数增多的现象。经过种植实验得出结论:十六倍体半夏生长期延长,抗逆性增强,夏季基本不倒苗,并且块茎个体均一,品质较好,总体产量增加。对十六倍体半夏块茎进行测定比较后发现,十六倍体半夏符合药典中规定的各项定性和定量指标规定。由此可见,通过半夏的单细胞进行多倍体的诱导是一种可行的方案,并且诱导得到的十六倍体为抗性增强、产量提高,且符合药典规定的半夏新品系。     

甜叶菊同源四倍体离体诱导及鉴定

用不同浓度秋水仙素溶液处理甜叶菊不定芽,诱导同源四倍体,并进行解剖学、染色体鉴定和流式细胞仪鉴定倍性。结果表明:(1)用0.20%的秋水仙素溶液浸泡甜叶菊不定芽12h,同源四倍体诱导率最高,可达32.14%。(2)同源四倍体植株与二倍体(对照)相比,其气孔、叶片等均表现巨大性,且叶片变厚、叶色浓绿、叶片皱缩。(3)对照植株染色体2n=2x=22,四倍体植株染色体2n=4x=44;流式细胞仪倍性鉴定结果显示,对照DNA相对含量为100,四倍体DNA相对含量为200。(4)该研究共鉴定出48株甜叶菊同源四倍体植株,为进行倍性植株的诱导奠定了技术基础,为进一步开展甜叶菊同源四倍体新品种的选育提供了实验材料。     

太子参快繁技术的优化及同源四倍体的诱导与鉴定

以太子参〔Pseudostellaria heterophylla(M iq.)Pax〕二倍体组培苗为实验材料,运用正交实验设计和组织培养方法,对太子参试管苗快速繁殖技术进行优化,并进行了同源四倍体的诱导与鉴定。结果表明,太子参最佳的繁殖培养基为含1.0 mg.L-16-BA和0.2 mg.L-1NAA的MS培养基;最佳生根培养基为含0.1 mg.L-1IAA、0.2mg.L-1NAA和1.0 mg.L-1ABT或0.2 mg.L-1IAA、0.2 mg.L-1NAA和1.0 mg.L-1ABT的1/2MS培养基。诱导同源四倍体的最佳处理方法为:用0.2%秋水仙素处理22 h或用0.3%秋水仙素处理12 h,所诱导的同源泉四倍体染色体数目为2n=4x=64条。     

野生南荻与芒杂种多倍体诱导研究

用不同浓度秋水仙素处理野生南荻×芒(Miscanthus lutarioriparia×Miscanthus sinensis)远缘杂交后代以诱导产生多倍体,并对变异株进行形态学和细胞学鉴定,以期获得稳定的四倍体植株并分析其生理特性。结果表明:(1)采用秋水仙素加入培养基处理法和秋水仙素溶液浸泡处理法都可获得一定频率的多倍体植株;胚性愈伤组织以0.2%秋水仙素浸泡处理48h的诱变效果较好,四倍体诱导率达8.7%;芽在0.05%秋水仙素培养基中处理15d较好,四倍体诱导率达10.6%;生根苗在0.1%秋水仙素培养基中处理10d较好,四倍体诱导率达11.1%。(2)经体细胞染色体计数,加倍植株染色体数为2n=4x=76,对照植株的染色体数目为2n=2x=38。(3)生长2年的多倍体植株形态、叶片大小、茎粗、茎壁厚、节间等性状表现出巨大性和超亲优势。     

秋水仙素诱导牛蒡多倍体

用种子处理法,以秋水仙素作为诱导剂,获得了牛蒡的同源四倍体(2n=4x=72),同时还得到了一些八倍体(2n=8x=144)和非整倍体植株.     

何首乌同源四倍体的诱导及生理指标的测定

用秋水仙素诱导何首乌(Polygonum multiflorumThunb.)同源多倍体的最佳条件为用0.2%秋水仙素溶液处理其试管苗幼嫩顶芽36 h,并接种于含2.0 mg.L-16-BA和0.2 mg.L-1IAA的MS启动培养基上。对诱导出的多倍体株系进行染色体计数分析,结果表明,诱导产生的多倍体均为四倍体,染色体基数2n=4x=44。生理指标测定结果表明,何首乌同源四倍体株系试管苗的超氧化物歧化酶(SOD)、过氧化物酶(POD)和黄嘌呤氧化酶(APX)活性及叶绿素含量等指标均高于二倍体株系。     

秋水仙素对草莓离体叶片再生和多倍体诱导的影响

以草莓(Fragaria×ananassa Duch.)栽培品种'雪蜜'(2n=8X=56)的离体叶片为外植体,研究了不同浓度秋水仙素对愈伤组织诱导率、不定芽再生率以及多倍体植株诱导的影响,并采用流式细胞仪对多倍体植株的倍性进行鉴定.结果显示,用质量体积分数0.1%、0.3%、0.5%和0.7%的秋水仙素浸泡2、4和6 d,草莓离体叶片均能诱导出愈伤组织和不定芽,但随秋水仙素浓度的提高和处理时间的延长,愈伤组织诱导率和不定芽再生率均显著下降.用不同浓度秋水仙素处理均能产生多倍体植株,倍性为9X、10X、11X、12X、14X和16X;随秋水仙素浓度的提高,多倍体诱导率呈现先上升后下降的变化趋势.用质量体积分数0.3%秋水仙素浸泡处理4 d是最佳的草莓离体叶片诱导方法,不定芽再生率达到40.5%,多倍体诱导率为100.0%,并且诱导产生出16X的植株.     

龙珠果茎段离体培养和组培苗耐盐性分析

为建立龙珠果(Passiflora foetida)的快繁再生体系,以实生苗茎段为外植体,研究了植物生长调节剂对丛生芽诱导、壮苗生根的影响,同时对组培苗的耐盐性进行研究。结果表明,MS+6-BA 0.5 mg/L+NAA 0.05 mg/L培养基有利于诱导丛生芽并促进芽的生长;MS+6-BA 3.0 mg/L+NAA 0.3 mg/L培养基有利于诱导愈伤组织;1/2 MS+IBA 0.2 mg/L培养基适合小芽壮苗生根。组培苗移栽至泥炭土∶蛭石∶珍珠岩(2∶1∶1)的基质中,成活率可达92.6%,且植株生长良好。0～200 mmol/L NaCl处理的组培苗生长不受影响;超过200 mmol/L NaCl处理,植株出现矮化、叶片萎蔫、变黄等现象。随NaCl浓度升高,叶片的SOD活性逐渐升高,POD、CAT和APX活性则呈先升高后降低的趋势。这为龙珠果的种苗繁育、海滨生态修复提供了技术支持。     

广藿香叶盘法高效再生体系主要影响因素的研究

广藿香是重要的芳香药用植物,利用基因工程技术对广藿香进行品种改良,需要建立一个高效的广藿香植株再生体系。该研究以广藿香无菌苗叶片为材料,将叶盘外植体分别置于不同条件下培养,观察、统计其再生植株的数量及生长状况。通过研究15~50 d的苗龄、第2~4节上的叶及培养基中2,4-D、NAA、BA和KT的浓度和配比等因素对广藿香叶盘再生植株的影响,在此基础上优化培养条件,建立广藿香高效再生体系。结果表明:广藿香无菌苗的苗龄、叶片在茎上的着生位置以及培养基中的植物生长调节物质浓度和配比都对广藿香的植株再生有显著影响;优化培养条件为以培养30 d的广藿香无菌苗顶芽下第2对展开叶片切割的叶盘为外植体,在含0.1 mg·L-1NAA和0.5 mg·L-1BA的MS培养基中培养28 d,叶盘的不定芽发生频率达到100%,单个叶盘的平均再生芽数为96.5个,经生根培养及温室炼苗,再生植株的移栽成活率达到96%。广藿香叶盘植株再生体系的建立为其基因转化研究及优良品种的快速繁育奠定了基础。     

紫茉莉不定芽诱导和植株再生

紫茉莉(Mirabilis jalapa)观赏价值较高,是一种重要的污染修复植物.组织培养技术为植物品种改良和选育的重要途径,但紫茉莉离体快繁方面的研究尚未见有相关报道.该研究以紫茉莉叶片和茎段为外植体,通过观察和统计外植体愈伤组织和不定芽的诱导情况,分析不同植物生长物质对紫茉莉植株再生的影响.结果表明:紫茉莉带芽茎段比较适合丛生芽的诱导,当带芽茎段在MS+1.0 mg·L-16-BA+1.5 mg·L-1 KT+1.0 mg·L-1 NAA+0.05 mg·L-1 TDZ培养基中培养时,不定芽的增殖系数较高.无论是MS或1/2MS培养基,都可诱导不定根的产生,其中生根效果较好的培养基为1/2 MS+0.5 mg·L-1 NAA.该研究结果探索了紫茉莉组织培养的最适条件,根据愈伤组织诱导率和不定芽的增殖系数筛选出了适宜不定芽诱导的培养基类型,根据不定芽生根情况确定了最佳的生根诱导培养基,为建立紫茉莉高效稳定的再生和遗传转化体系奠定了基础.     

Effect of polyploidy induction on biomass and ginsenoside accumulations in adventitious roots of ginseng

Adventitious roots ofPanax ginseng C.A. Meyer (a natural tetraploid) were treated with 50 or 100 mg L^-1 colchicine for 12, 24,36, 48, or 60 h to induce polyploid (octoploid) roots. The largest number of octoploid roots was obtained with a 100 mg L^-1 colchicine treatment over 60 h. To verify that ginsenoside was being accumulated in the developing tissues, the tetraploid (control) and octoploid roots were cultured for 40 d in Murashige and Skoog media that lacked NH₄NO₃ but was supplemented with 2 mg L^-1 naphthaleneacetic acid and 50 g L_-1 sucrose. Levels of fresh and dry biomass were greater in the octoploid roots. Although total ginsenoside and Rb-group ginsenoside contents were less in the octoploid roots than in the tetraploids, the former had a higher amount of Rg-group ginsenosides (especially Rg1). These results demonstrate the benefit that polyploid adventitious roots provide in enhancing the production of secondary metabolites in ginseng.     

Plant regeneration via organogenesis from adventitious bud explants of a medicinal herb species, <Emphasis Type="Italic">Polygonatum cyrtonema</Emphasis>

Summary Explants derived from adventitious buds, rhizomes, stems, and leaves of a medicinal plant, Polygonatum cyrtonema, were studied for plantlet regeneration, and only adventitious bud explants were able to be regenerated into plantlets. Regeneration was also accompanied by the formation of rhizome-like tissue, the medicinal portion of the plant. The optimum hormone combination for plantlet regenertion was 4.44 μM benzyladenine plus 2.26 μM 2,4-dichlorophenoxyacetic acid, at which new adventitious buds were obtained from 96.6% of the adventitious bud explants, with an average of 5.2 buds per explant. The best medium for root induction was half-strength Murashige and Skoog medium with 4.57 μM α-naphthaleneacetic acid, as 92% of regenerated buds rooted. Regenerated plantlets were successfully transferred to a greenhouse with 86% survival. Histological observation indicated that new adventitious buds originated from the superficial meristematic cell cluster of the granular callus induced from adventitious bud explants via organogenesis.     

High-frequency plant regeneration from leaf-disc cultures of Jatropha curcas L.: an important biodiesel plant

A simple, high-frequency and reproducible protocol for induction of adventitious shoot buds and plant regeneration from leaf-disc cultures of Jatropha curcas L. has been developed. Adventitious shoot buds were induced from very young leaf explants of in vitro germinated seedlings as well as mature field-grown plants cultured on Murashige and Skoog’s (MS) medium supplemented with thidiazuron (TDZ) (2.27 μM), 6-benzylaminopurine (BA) (2.22 μM) and indole-3-butyric acid (IBA) (0.49 μM). The presence of TDZ in the induction medium has greater influence on the induction of adventitious shoot buds, whereas BA in the absence of TDZ promoted callus induction rather than shoot buds. Induced shoot buds were multiplied and elongated into shoots following transfer to the MS medium supplemented with BA (4.44 μM), kinetin (Kn) (2.33 μM), indole-3-acetic acid (IAA) (1.43 μM), and gibberellic acid (GA₃) (0.72 μM). Well-developed shoots were rooted on MS medium supplemented with IBA (0.5 μM) after 30 days. Regenerated plants after 2 months of acclimatization were successfully transferred to the field without visible morphological variation. This protocol might find use in mass production of true-to-type plants and in production of transgenic plants through Agrobacterium/biolistic-mediated transformation.     

Adventitious shoot formation in cultures of 30-year-old Larix decidua,L. leptolepis,L. eurolepis,and L. laricina trees

Primordial shoot explants excised from buds of one Larix decidua tree, about 30 years old, produced more adventitious buds, elongating into shoots, when grown on half strength Litvay medium than when grown on other basal media. Thidiazuron and N₆-benzyladenine (BA) were equally effective in adventitious bud induction. In a comparative study of 30-year-old L. decidua, L. leptolepis, L. eurolepis, and L. laricina trees, explants from L. eurolepis and L. decidua produced a high number of cultures with adventitious buds that elongated into shoots; those from L. leptolepis were less productive, and those from L. laricina failed to form adventitious buds. The highest response was obtained with material collected in August and September, and in March and April; the lowest response occurred in explants from the October collection.     

两种栽培型广藿香内生真菌群落组成变化

为探索内生真菌与广藿香互作间对宿主活性成分形成机制的影响,该研究以成分差异较大的牌香和湛香为对象,采用传统形态学方法对所获菌株归类,通过真菌通用引物ITS1/ITS4扩增菌株rDNA-ITS序列,鉴定其分类地位并研究其多样性。结果表明：(1)用PDA和LBA培养基对苗期、分枝期和成株期广藿香茎叶组织块进行内生真菌分离,共获得3 070株菌株,其中牌香(PX)分离出1 624株,鉴定出1 319株,分属于36属;湛香(ZX)分离出1 446株,鉴定出994株,分属于33属。牌香分离出7种特有内生真菌,分别为香柱菌(Epichloe typhina)、盘长孢状刺盘孢菌(Colletotrichum gloeosporioides)、座腔孢菌(Botryosphaeria sp.)、丝核菌(Rhizoctonia sp.)及截盘多毛孢菌(Truncatella sp.),并首次分离到疫霉菌(Phytophthora sp.)和指疫霉菌(Sclerophthora sp.),这2种菌属于卵菌门内生菌。湛香分离出拟青霉菌(Paecilomyces sp.)和尾孢菌(Cercospora sp.)...     

Micropropagation of wild beet (Beta maritima) from inflorescence pieces

In order to investigate the regeneration of wild beet (Beta maritima) from inflorescence pieces, the effects of growth regulator, genotype, explant source and stage of plant development on adventitious shoot formation and rooting in vitro and subsequent transplanting in the glasshouse were tested. Inflorescence tips produced more adventitious shoots than sub-apical segments and the best micropropagation was achieved on a Murashige and Skoog (MS) medium supplemented with 1.0 mg l^–1 BAP. Addition of auxin was not beneficial. The induction rate of adventitious shoots was genotype-dependent and influenced by the stage of plant development. Adventitious shoots were produced from the base of the flower buds, i.e. from the receptacle, not from axils or stalks and only a few buds on inflorescence tip explants produced adventitious shoots. Rooting was increased by using a MS medium with 3% sucrose supplemented with 1.0 mg l^–1 NAA. There was no variation in leaf morphology of the transplants. This work shows that inflorescence tips can be used successfully as explants for in vitro multiplication of sugar beet and wild beet.Abbreviations BAP benzylaminopurine - IBA indole-3-butyric acid - GA₃ gibberellic acid - MS Murashige and Skoog medium - NAA naphthaleneacetic acid Author for correspondence     

An investigation of morphogenesis within the genus Trifolium

Morphogenic responses within the genus Trifolium were investigated by culturing various explants from seedlings of 72 species. Seedlings from 32 species produced callus alone, 40 produced adventitious shoots and/or roots, of which 25 species produced only shoots and 7 species formed only roots. Seedlings within each species also varied in their response to culture. The section of these seedlings most likely to produce adventitious shoots was the original shoot with the remnants of the surrounding hypocotyl and cotyledons, followed by the excised cotyledons themselves.Inter- and intra-varietal variation was observed in T. repens. Genotypes that produced adventitious buds were selected and crossed. An improvement in the proportion of the population capable of morphogenesis was observed in one cultivar.