瘤蕨孢子组织培养与快速繁殖(简报)

以瘤蕨的孢子为外植体进行组织培养研究。结果表明,瘤蕨的成熟孢子在1/2 MS培养基上萌发速度最快,萌发率最高;瘤蕨的原叶体在1/2MS + 1.0 mg/L 6-BA + 0.5 mg/L NAA培养基上增殖速度较快,但增殖过程中不能形成孢子体;在培养基1/2MS +1.0 m/L AgNO3 + 0.05 mg/L NAA上成功诱导出孢子体;孢子体在培养基1/2MS + 0.5 mg/L IAA上可进行分株扩增,最终获得大量孢子无菌苗。     

同形鳞毛蕨成精子囊素系统的初步研究

研究了同形鳞毛蕨成精子囊素对该种和水蕨孢子萌发和配子体发育的影响,结果表明：同形鳞毛蕨配子体能产生成精子囊素,该成精子囊素能抑制同种孢子的萌发,抑制作用随配子体成熟度的增加而增强;同形鳞毛蕨成精子囊素还可促进同种孢子发育为雄配子体;光照条件下,同形鳞毛蕨成精子囊素对水蕨孢子萌发和配子体发育影响不大,黑暗条件下,同形鳞毛蕨成精子囊素能显著的促使水蕨孢子提早萌发,但都不影响其孢子最终萌发率和配子体的性别分化,表明同形鳞毛蕨和水蕨的成精子囊素不属于同一系统。     

孢子大小和培养基对水蕨孢子萌发及配子体性别分化的影响

利用MS培养基、改良Knop’s培养基、自来水和蒸馏水分别培养水蕨中等大小孢子,同时利用改良Knop’s培养基培养不同大小的水蕨孢子,观察记录不同条件下水蕨孢子萌发和性别分化情况。实验表明,二级孢子（赤道轴120～140μm）萌发率最高;一级孢子（赤道轴〉140μm）萌发最有利于使水蕨发育为两性配子体,三级孢子（赤道轴〈120μm）萌发最有利于使水蕨发育为雄配子体;MS培养基和改良Knop’s培养基相对于自来水和蒸馏水有利于水蕨孢子萌发;各培养基中水蕨两性配子体比率排序是MS培养基〉改良Knop’s培养基〉自来水〉蒸馏水,而雄配子体比率排序与之相反。此结果为水蕨的引种保护、人工繁育和分子生物学研究提供理论依据。     

银粉背蕨配子体及幼孢子体发育的研究

银粉背蕨是一种小型观赏蕨类植物,但目前我国对该蕨的研究还不够成熟。本文利用改良Knop's培养基和腐殖土培养银粉背蕨的孢子,观察其配子体及幼孢子体形态发育特征,并研究了其配子体发育的最适培养基pH值。研究结果显示：（1）银粉背蕨孢子黄褐色,具三裂缝,极面观三角圆形,赤道面观为近半圆形,孢子具网状纹饰;孢子萌发为书带蕨型;原叶体发育为水蕨型;颈卵器和精子器为薄囊蕨型;成熟原叶体为对称的心脏形,不具毛状体;上述特征为银粉背蕨孢子和配子体发育的稳定特征。（2）培养基pH值在7.0~9.0时随着碱性的增强,银粉背蕨孢子萌发和配子体生长发育速度逐渐增加。（3）利用腐殖土培养银粉背蕨孢子,7~8周可发育成幼叶,成苗率达90%,成苗健壮,根系发达,是扩繁银粉背蕨的适宜方式。本文为资源植物银粉背蕨人工繁殖和演化研究提供科学依据。     

Establishment of a Tissue Culture and Rapid Propagation System of Dryopteris fragrans

香鳞毛蕨(Dryopteris fragrans)是一种多年生野生草本蕨类植物, 具有抗氧化、抑菌、抗银屑病和抗肿瘤等多种功效。香鳞毛蕨的野生资源匮乏, 通过组织培养建立其人工再生体系, 是保护香鳞毛蕨资源可持续利用的有效途径。通过对香鳞毛蕨孢子的无菌培养, 比较分析不同因素对原叶体增殖、孢子体诱导、愈伤组织诱导与增殖、丛生芽分化及生根的影响, 建立了快速繁殖体系, 为香鳞毛蕨规模化生产奠定基础。研究结果表明, 当培养基组分为1/2MS时, 原叶体生长状态较好, 颜色翠绿, 增殖倍数最高可达5.67±0.59, 此时有大量幼孢子体产生, 孢子体诱导率为(37.50±2.04)%; 愈伤组织诱导最佳培养基为1/2MS+2.0 mg·L^-1 6-BA+1.0 mg·L^-12,4-D, 诱导率可达(96.67%±5.77)%; 愈伤组织增殖最佳培养基为1/2MS+ 1.0 mg·L^-1 6-BA+0.5 mg·L^-1 2,4-D, 增殖倍数达13.30; 颗粒状愈伤组织在1/2MS培养基中生长出大量丛生芽, 诱导率为(53.33±3.33)%; 1/2MS+0.2 mg·L^-1 NAA培养基促进幼孢子体苗生根, 移栽成活率约为60%。     

香鳞毛蕨的组织培养和快速繁殖体系构建

香鳞毛蕨(Dryopteris fragrans)是一种多年生野生草本蕨类植物, 具有抗氧化、抑菌、抗银屑病和抗肿瘤等多种功效。香鳞毛蕨的野生资源匮乏, 通过组织培养建立其人工再生体系, 是保护香鳞毛蕨资源可持续利用的有效途径。通过对香鳞毛蕨孢子的无菌培养, 比较分析不同因素对原叶体增殖、孢子体诱导、愈伤组织诱导与增殖、丛生芽分化及生根的影响, 建立了快速繁殖体系, 为香鳞毛蕨规模化生产奠定基础。研究结果表明, 当培养基组分为1/2MS时, 原叶体生长状态较好, 颜色翠绿, 增殖倍数最高可达5.67±0.59, 此时有大量幼孢子体产生, 孢子体诱导率为(37.50±2.04)%; 愈伤组织诱导最佳培养基为1/2MS+2.0 mg·L^-1 6-BA+1.0 mg·L^-12,4-D, 诱导率可达(96.67%±5.77)%; 愈伤组织增殖最佳培养基为1/2MS+ 1.0 mg·L^-1 6-BA+0.5 mg·L^-1 2,4-D, 增殖倍数达13.30; 颗粒状愈伤组织在1/2MS培养基中生长出大量丛生芽, 诱导率为(53.33±3.33)%; 1/2MS+0.2 mg·L^-1 NAA培养基促进幼孢子体苗生根, 移栽成活率约为60%。     

假瘤蕨属植物配子体与幼孢子体发育的比较形态学研究

配子体及幼孢子体发育过程对蕨类植物的系统学研究具有重要意义,但在假瘤蕨属植物中较少报道.本研究比较了3种假瘤蕨属植物的配子体及幼孢子体发育过程:孢子均为单裂缝,萌发类型为书带蕨型.原叶体发育为槲蕨型.丝状体2～6细胞,成熟配子体心形,中肋明显加厚.配子体两性,在播种后48 ～ 55 d产生精子器,之后15 ～ 18 d产生颈卵器.播种后80 ～ 100 d,形成胚胎,后者分化出第一叶、第一根和茎端,发育为幼孢子体.配子体边缘分布有单细胞毛状体,配子体腹面分布有单列多细胞的毛状体,以中肋处最多,围绕并保护胚胎和幼孢子体.本属3个种的配子体和幼孢子体,在孢子体积、萌发时间、丝状体和成熟配子体特征以及性器产生时间等方面存在差异.土培条件下的配子体发育不同步,即配子体分批发育,原因为配子体的营养繁殖或孢子萌发不整齐.     

扇蕨配子体发育与濒危机制的探讨

采用改良Knop’s固体培养基、原生境腐殖质土和红壤分别培养扇蕨（Neocheiropteris palmatopedate）孢子,光学显微镜及解剖镜下观察记录其孢子萌发及配子体发育过程,比较了3种培养方法对其配子体发育和有性繁殖的影响,并在此基础上探讨了扇蕨的濒危原因。结果表明：成熟孢子黄褐色,赤道面观为豆形,极面观椭圆形,单裂缝。孢子萌发类型为书带蕨型,原叶体发育为槲蕨型。成熟原叶体呈心脏形。毛状体在原叶体阶段出现。有性生殖周期长及配子体发育成幼孢子体的百分率低是扇蕨在配子体世代的主要濒危原因。此外,红壤固有的理化性质导致扇蕨配子体发育极其缓慢、精子器和颈卵器发生的时间间隔过长使其不能受精产生孢子体。原生植被遭受破坏引起的林下腐殖质土消失、红壤裸露,加剧了扇蕨的濒危。     

液体培养基条件下乌毛蕨配子体发育的研究

在液体培养基条件下对中国产乌毛蕨(Blechnum orientaleL.)配子体发育进行了观察。结果表明:乌毛蕨的孢子为两面体型,两侧对称,极面观为椭圆形,赤道面观为半圆形或豆形,周壁具脊状褶皱;孢子萌发为书带蕨型;原叶体发育为三叉蕨型,成熟原叶体为对称的心脏形。经比较分析,蕨类植物孢子繁殖时采用混合土培养较适宜;液体培养基和混合土在研究蕨类植物配子体发育时同样具有可行性。孢子形态和配子体发育的观察结果表明,乌毛蕨是蕨类植物中较进化的种类;乌毛蕨科与鳞毛蕨科亲缘关系较近。     

鳞毛蕨科3种植物配子体发育的研究

利用光学显微镜详细观察鳞毛蕨科(Dryopteridaceae)3属3种植物,即耳蕨属的棕鳞耳蕨(Polystichum polyblepharum)、鳞毛蕨属的黑足鳞毛蕨(Dryopteris fuscipes)和鞭叶蕨属的鞭叶蕨(Cyrtomidictyum lepidocaulon)配子体发育过程,记录配子体各发育阶段的特征。结果表明:棕鳞耳蕨、黑足鳞毛蕨和鞭叶蕨的孢子都为单裂缝,孢子萌发类型为书带蕨型,配子体发育类型为三叉蕨型,性器官为薄囊蕨型,成熟原叶体为对称的心脏形,具毛状体;3属的属间差异包括孢子颜色、孢子纹饰、萌发时间、丝状体长度、片状体形状、毛状体和假根等特征,上述特征在属间存在交叉现象,因此,孢子形态和配子体发育特征不能作为区分耳蕨属、鳞毛蕨属和鞭叶蕨属的形态依据。     

Biological and nutritional aspects involved in fern multiplication

Gametophytes of several species of ferns were mechanically triturated and the resulting homogenates cultured in vitro for propagation purposes. Differences in the time period from spore culture to sporophyte development were perceivable between species. For those species with a fast life cycle and high sporophyte production such as Woodwardia virginica and Dryopteris affinis sp. affinis, homogenization of gametophytes can be considered to be excellent method for propagation, yielding hundreds of sporophytes in a short period of time. Sporophyte formation was inhibited in O. regalis by the succesive application of homogenization to gametophytes regenerated by this technique. The effect of the culture medium composition on fern production was also studied in O. regalis and P. ensiformis gametophytes. In these species, sporophyte formation increased when the gametophytes were cultured in a medium containing water+0.7% agar. Addition of sucrose inhibited gametophyte development and induced their necrosis. The 1/2 dilution of Murashige and Skoog basal medium, without sucrose, favoured leaf expansion in P. ensiformis sporophytes. This revised version was published online in June 2006 with corrections to the Cover Date.     

Practical methodology for gametophyte proliferation and sporophyte production in green penny fern (Lemmaphyllum microphyllum C. Presl) using mechanical fragmentation

Propagation of gametophytes and sporophytes using mechanical fragmentation has been considered a suitable method for mass production of ferns. This study aimed to develop a practical propagation method for Lemmaphyllum microphyllum C. Presl, which is a fern of significant ornamental and medicinal value. Gametophytes were obtained through in vitro spore germination and used for propagation experiments. The gametophyte was mechanically fragmented using a scalpel into small fragments, which were then used to investigate gametophyte proliferation. In addition, the gametophyte was fragmented using a blender and then used to study sporophyte formation. Optimal proliferation conditions of the gametophyte were determined using Murashige and Skoog (MS) basal medium (double-, full-, half-, quarter-strength), Knop medium, and medium components (sucrose, nitrogen sources, activated charcoal), at various concentrations. The fresh weight of the gametophyte was 14-fold higher than that of gametophytes (300 mg) used as culture material, when cultured on double-strength MS. Moreover, 1 g of the gametophyte fragmented in 25 mL of distilled water formed more than 430 sporophytes in a soil mixture in an area of 7.5 cm². The sporophytes were successfully cultivated in the greenhouse after acclimation. A large-scale production method for L. microphyllum that can be easily implemented in a fern production farm is outlined.

     

鸟巢蕨孢子繁殖技术研究

研究了鸟巢蕨孢子无菌播种和常规播种两种繁殖方法。结果表明,鸟巢蕨无菌播种中孢子萌发率最高可达66.7%,原叶体在固体培养基上培养不能诱导出孢子体,而需经过振荡培养后才能诱导出孢子体;常规播种法更容易诱导出孢子体,每盆播0.02 g孢子时,每克孢子可产生孢子体4 000株以上,比无菌播种操作简便,成本低。因此,孢子常规播种法更适合于鸟巢蕨规模化生产。     

阔鳞鳞毛蕨孢子的无菌繁殖

以阔鳞鳞毛蕨（Dryopteris championii）成熟孢子为外植体,研究了不同植物生长调节剂及浓度对其孢子萌发、愈伤组织诱导、丛生芽分化及生根的影响。结果表明：1/2MS＋2%蔗糖为孢子萌发最适培养基,20 d后萌发率达65%;诱导愈伤组织的最适培养基为MS＋KT 0.5 mg.L-1＋2,4-D 1 mg.L-1,诱导率达39.8%,愈伤组织为绿色颗粒状;颗粒状愈伤组织在MS＋KT0.2 mg.L-1的培养基中生长出大量丛生芽,转化率可达66.3%;MS＋IAA 0.2 mg.L-1＋NAA 0.2 mg.L-1培养基可有效促进幼孢子体苗生根。     

Sporophyte and gametophyte development of Platycerium coronarium (Koenig) Desv. and P. grande (Fee) C. Presl. (Polypodiaceae) through in vitro propagation

The sporophyte and gametophyte development of Platycerium coronarium and P. grande were compared through ex situ propagation using in vitro culture technique and under greenhouse and field conditions.The morphology of the sporophyte and gametophyte, type of spore germination and prothallial development of P. coronarium and P. grande were documented. Gametophytes of P. coronarium and P. grande were cultured in vitro using different media. The gametophytes were then transferred and potted in sterile chopped Cyathea spp. (anonotong) roots and garden soil for sporophyte formation. Sporophytes (plantlets) of the two Platycerium species were attached on the slabs of anonotong and on branches and trunks of Swietenia macrophylla (mahogany) under greenhouse and field conditions.Sporophyte morphology of P. coronarium and P. grande varies but not their gametophyte morphology. P. coronarium and P. grande exhibited rapid spore germination and gametophyte development in both spore culture medium and Knudson C culture medium containing 2% glucose. Gametophytes of P. coronarium and P. grande transferred to potting medium produced more number of sporophytes while the gametophytes inside the culture media did not produce sporophytes. Sporophytes of P. grande attached on mahogany branches produced more number of leaves with bigger leaf area than those attached on anonotong slabs. Likewise, sporophytes of P. coronarium attached on mahogany branches and anonotong slabs did not develop new leaves during two weeks monitoring and are still in a period of adjustment to its environment. Sporophytes of P. grande grown or attached on the trunk of mahogany trees in the field and under shaded environment favored their growth.     

A methodology for large-scale Athyrium sheareri gametophyte proliferation and sporophyte production using tissue culture

Tissue culture methods using gametophytes are considered the easiest ways to mass-produce fern sporophytes. The aim of this study was to develop a practical propagation method for the ornamental fern, Athyrium sheareri. The gametophytes obtained from in vitro spore germination were used as experimental materials. We used the chopping method to investigate the culturing conditions for proliferating gametophytes and the blending method for evaluating the mass production of sporophytes in mixed soil. Gametophyte proliferation was determined via Knop medium, various concentrations of Murashige and Skoog (MS) basal medium (1, 1/2, 1/4), and media components (sucrose, nitrogen source, and activated charcoal). The fresh weight of the gametophytes increased by more than 24-fold in 1/2 MS medium. In addition, 1 g of gametophyte could produce a maximum of 255.3 sporophytes in a mixed soil of 7.5 cm² area. Treating gametophytes with exogenous plant growth regulators promoted the formation and growth of sporophytes. The cultivated young sporophytes were acclimated and successfully grown in greenhouses. We developed a mass production protocol for A. sheareri sporophytes suitable for field application, which is expected to have commercial value.

     

2种培养基下紫萁配子体发育及孢子体形成的研究

采用光镜技术对不同培养基下紫萁配子体发育和孢子体形成进行了研究.结果表明:(1)紫萁孢子绿色,四面体形,具三裂缝,孢子两极萌发分别产生假根和原叶体细胞,原叶体细胞形成球形体,球形体产生分生细胞发育为片状体和原叶体.(2)原叶体可为雄性、雌性或两性;雄原叶体较小,其两翼基部产生多数精子器;雌原叶体大,具有明显的中肋,在中...     

中华鳞毛蕨配子体发育过程的观察

Morphological variation during the gametophyte development process of Dryopteris chinensis ( Bak.) Koidz. was observed. The results show that the development process of D. chinensis can be divided into spore germination, filament formation, plate formation, prothallus formation, sexual organ formation and apogamety. The spores are bilaterally symmetric, monolete, surface with ridge fold ornamentation, elliptical in polar view, and approximately semicircle-shaped in equatorial view. The spore germination type is of Vittaria-type;the filaments with a length of 3-7 cells, not branched or occasionally branched, with single or double row cells; mature prothallus is symmetrically cordate, and the development type of prothallus is of Aspidium-type, with long unicellular clavate trichomes distributed on the surface and in the margin;with antheridium but archegonium is not observed, belongs to apogamety. One prothallium of D. chinensis produces one embryo, and young embryo can be generated within one month after the formation of antheridium; there are a lot of unicellular trichomes and some multicellular trichomes on the young embryo of sporophyte.     

粗茎鳞毛蕨孢子萌发研究

以经过3年低温储藏的粗茎鳞毛蕨孢子为实验材料,从孢子离心、孢子消毒、培养基种类、光质等4方面对孢子萌发进行研究,结果表明:在离心转数≤14 000 r.min-1、离心时间≤30 min条件下,离心处理对孢子萌发基本无影响;对孢子进行1%NaClO水溶液浸泡处理20～30 min为最佳消毒条件;改良Knop’s培养基为最佳孢子萌发培养基;黑暗条件下孢子不能萌发,但是黑暗处理能够明显提高孢子萌发整齐性;红光比白光能促进孢子提早萌发1 d左右,但对提高萌发率效果不显著。