ABA,NAA诱导水稻胚性愈伤组织的研究

ＡＢＡ和ＮＡＡ联合使用能有效地诱导水稻原生质体再生的愈伤组织向胚性发展。通过液体浅层培养由原生质体得到的愈伤性发展。通过液体浅层培养由原生质体得到的愈伤组织,在含ＡＢＡ和ＮＡＡ的Ｎ８培养基上培养一段时间,可以诱导原来呈非胚性状态的愈伤组织形成胚性愈伤组织,并在含ＺＴ的Ｎ６分化培养基上产生绿点。通过对这两种愈伤组织的生化分析,表明二者在游离氨基酸、ＤＮＡ、ＲＮＡ、核酸及蛋白质含量等方面,特别是ＳＤＳ     

水稻原生质体再生小植株

水稻的原生质体培养一直是引人注意的问题,多年来进展不大。近年来Fujimura等Coulibaly等Yamada等分别由水稻盾片和未成熟胚愈伤组织原生质体;胚芽鞘幼嫩愈伤组织的原生质体和水稻雄性不育系细胞游离原生质体得到再生植株。我们用水稻(Oryzasativa)农虎6号种子胚诱导愈伤组织制备原生质体,也培养得到再生小植株,结果简报如下:     

小叶旱烟(Nicotiana rustica)茎愈伤组织的原生质体再生成完整植株的研究

用纤维素酶成功地从小叶旱烟茎愈伤组织中分离出了大量具有活力的原生质体。在所确定的培养条件下,由原生质体培养出了愈伤组织,并发育成完整植株。同时,对IAA、α-NAA和激动素在原生质体再生成完整植株中的作用进行了研究。     

影响谷子原生质体看护培养的一些因素

研究了谷子原生质体看护培养中一些问题。结果表明：不同种植物愈伤组织做看护细胞对谷子原生质体培养植板率有不同的影响。用光头稗草愈伤组织对谷子胚性、非胚性或中间型愈伤组织的原生质体进行看护培养,以对胚性愈伤组织原生质体的效果最好。看护培养主要是作用于原生质体形成完整细胞后的生长发育,试验没有观察到明显的看护细胞数量效应。     

陆地棉原生质体高频率分裂及植株再生

取陆地棉品种（系）３１１８、９５５４和晋棉４号种子无菌苗的下胚轴诱导的愈伤组织,从中挑选具有分化能力的黄色颗粒状愈伤组织,建立胚性细胞悬浮培养系。以纤维素酶和离析软化酶组成的酶液,由细胞悬浮培养物游离原生质体。采用含低融点脂糖的Ｋ３基本培基包埋原生质体的培养方式,获得愈伤组织。以液体－固体－液体轮回培养法改良晋棉４号的细胞悬浮系,原生质体的植板率从２％左右提高到９％以上。在原生质体再生愈伤组织的继     

黑果枸杞原生质体培养及植株再生

该研究以黑果枸杞(Lycium ruthenicum)无菌苗为材料,建立了愈伤组织来源的原生质体再生体系,采用ISSR和FCM技术对再生植株进行了遗传稳定性分析。结果表明：(1)黑果枸杞叶片愈伤组织是产生原生质体的最好材料,在含0.5 mg·mL^-1甘露醇的酶液中,继代1次的叶片愈伤组织中原生质体产量为7.77×10⁶个·g^-1,活力为92%。(2)改良MS培养基 固体液体双层培养(MS₂ 固液双层)是培养原生质体的最好方式,培养10 d的原生质体分裂频率为45.9%,培养20 d的细胞团形成频率为22.9%。(3)在1.5 mg·mL^-1 6 BA+0.1 mg·mL^-1 IBA+MS培养基中,叶片愈伤组织产生的原生质体可分化获得再生植株。(4)ISSR分析显示,再生植株的平均遗传相似系数为0.88;FCM显示再生植株为二倍体,与亲本植株一致。该研究结果为进一步研究枸杞体细胞杂交技术转移野生植物抗逆遗传性状提供科学依据,为枸杞优良品种的选育奠定了基础。     

甘蔗原生质体的植物再生

从甘蔗嫩叶外植体诱导愈伤组织,经继代培养后,挑选胚性愈伤组织,转入ＭＳ３液体培养基,进行悬浮培养,当培养物分离出小粒状的细胞团,细胞变得小而圆时,用于分离原生质体,原生质体以琼脂糖固化的培养方式培养于ＭＲＰ１培养基中,由原生质体再生的愈伤组织有两种类型,挑选粒状,坚实的再和愈伤组织转移到Ｎ６分化培养基上,“新台糖１号“再生的愈伤组织,在含有ＫＴ０．５ｍｇ／Ｌ的培养基中,分化出绿芽并长成完整的植株。     

谷子胚性愈伤组织粘液提取物对原生质体培养的影响

将谷子胚性愈伤组织粘液提取物以谷子原生质体培养基中,其对原生质体培养的影响表明该提取物有助于原生质体形成的细胞壁;并且该类有粘液的愈伤的原生质体游离所需的酶液的愈伤组织的原生质体游离所需的酶液浓度低,处理时间短。由原生质休形成这完整细胞的数量在一定范围内与谷子原生质体培养的植板率相对应。通过增加形成完整细胞的数量较大幅度地提高原生持体培养的植板率。     

影响谷子原生质体看护培养的一些因素

研究了谷子原生质体看护培养中的一些问题。结果表明：不同种植物愈伤组织做看护细胞对谷子原生质体培养植板率有不同的影响。用光头种草愈伤组织对谷子胚性、非胚性或中间型意伤组织的原生质体进行看护培养,以对胚性意伤组织原生质体的效果最好。看护培养主要是作用于原生质体形成完整细胞后的生长发育。试验没有观察到明显的看护细胞数量效应。     

高粱原生质体培养再生植株

在高梁的组织培养方面,已从花药培养诱导愈伤组织再生植株,并通过茎尖培养建立了能产生大量绿苗的无性繁殖系。而在原生质体研究方面,虽然Brar DS等由未成熟胚诱导愈伤组织建立的悬浮细胞系分离原生质体,并得到了愈伤组织,但至今未见     

Plant Regeneration from Protoplast of Peucedanum praeruptorum Dunn

Calll were initiated from the seedling segment of Peucedanum praeruptorum Dunn and subcultured on the MS agar medium with 0.5 mg/L 2,4-D. Cell suspension culture with a lot of embryogenic cell clumps was obtained in liquid medium. Protoplasts were isolated from the cell clumps in enzyme mixture solution containing 1.5% Onozuka R-10, 0.3% Macerozyme R-10, 0.5% helicase, 5 mmol/L CaCl2 and 0.6 mol/L mannital, at pH 5.6 and shaking for 5- hours at 25℃. Helicase is necessary for isolation. After purified by washing, the protoplasts were cultured in liquid medium containing 1 mg/L 2,4-D +0.5 mg/L zeatin. First cell division was observed after four days. Large cell clumps were formed after thirty days. Microcalli of 1 mm in size was formed after about fifty days, and continued to grow on the MS solid medium containing 0.5 mg/L 2,4-D and 200 mg/L casein hydrolysate, and later differentiated into embryoids when transferred to MS agar medium with 0.1 mg/L zeatin. Eventually, embryoids developed into whole plantlets on the MS solid medium without phytohormones.     

埃斯基红豆草下胚轴愈伤组织原生质体的培养与植株再生

埃斯基红豆幼苗的下胚轴切段在附加２,４－Ｄ０．５ｍｇ／Ｌ,ＫＴ１ｍｇ／Ｌ的ＭＳ中形成胚性愈伤组织。来自１１－１３个月龄、继代６－１５天的愈伤组织的原生质体,在改良的Ｖ－ＫＭ液体培养基中可持续分裂形成细胞团,培养１０天时的分裂率和克隆率分别为６５．８８％和５３．３８％周后就可将将原生质体形成的小愈伤组织转于培养基上。原生质体在改良的Ｂ５液体培养基也可以分裂形成小愈伤组织,但分裂率低于Ｖ－ＫＭ。来自原     

Plant Regeneration from Cell Supension Derived Protoplasts of Heracleum moellendorffii

Heracleum moellendorffiz Hance is a herb belonging to Umbelliferae used in traditional medicine in China. The young stem-nodes were induced for callus formation on MS medium containing 1 mg/L 2,4-D. After subcultured for about five months, the embryogenic calli were used for cell suspension culture. The protoplasts were prepared from this suspension by digestion with enzyme mixture containing 1. 5% cellulase Onozuka R-10 +0. 3% macerozyme R-10 + 0. 5% snailase + 5 mmol CaCl2 + 0. 6 mol/L mannitol, at pH 5.8, and cultured in modified MS and modified N6 media with 0.3 % agarose. They divided after 3 days and developed into small cell colonies after about 2 weeks. From this time on, the glucose concentration in the culture media was decreased to 0. 2 mol/L,which led to futher growth of the colonies to small calf . After a period of proliferation on solid medium with 0. 5 mg/L 2,4-D, the calli were transferred to a medium with 0. 1 mg/L zeatin on which somatic embryos differentiated and developed to plantlets     

Preliminary studies on tissue culture and agrobacterium-medicated transformation of Brassica campestris ssp. chinensis]

The hypocotyls and cotyledons of the asepetic seedling of Brassica campestris ssp. chinensis L cv. Pudongaijiecai) were used as explants for tissue culture. Adventitious buds were differentiated on modified MS medium supplemented with TDZ 1-2 mg/L, NAA 0.2-1 mg/L and AgNO3 7.5 mg/L. The percentage of explants which formed buds of cotyledons was about 56%, and that of hypocotyls was about 37%. When the regenerated explants were transferred onto MS medium with 2 i.p. 5 mg/L and NAA 0.1 mg/L for two weeks, whole plantlets were obtained by culturing the regenerated shoots on 1/2 MS medium with NAA 0.1 mg/L. Agrobacterium tumefaciens strain (LBA 4404/PBI 121) carrying the GUS gene and Npt II gene was used for transformation. After 2 days of coculture, the hypocotyls and cotyledons were transferred onto regenerated medium containing CP 300 mg/L for bud formation. After 4-5 weeks, the differentiated buds were transferred onto selection medium with CP 200 mg/L and Km 10 mg/L for 1 month, then the green shoots were transferred onto the rooting medium containing Cef 100 mg/L and Km 20 mg/L. 4-5 weeks later, plantlets with Km resistance were obtained and some of them showed higher enzymatic activities of beta-glucuronidase than control ones.     

Plant Development from Isolated Microspores of Oryza sativa L. ssp. indica Derived from the Anther Culture of F2 Hybrids of Photoperiod-Sensitive Male-Sterile Rice

Microspores from a highly anther culturable rice line (Oryza sativa L. spp. indica) derived from the anther culture of F2 hybrids of photoperiod-sensitive male-sterile rice, after 7 days of low temperature treatment and another 7 days of preculture within anthers, were isolated mmechanically. They were cultured in Ne medium containing 3% manitol, 6% sucrose, 5 g/L inositol, 100 mg/L serine, 800 mg/L glutamine, 1 000 mg/L L-proline, 10% (V/V) coconut milk and 2 mg/L, 2,4-D, and 1 mg/L kinetin. After 5 days, microspores initiated first division and subsequently developed into multicellular pollens and calli. Green plant could be recovered when compact calli were transferred onto agar-solidified MS medium containing 3% sucrose, 0.5 mg/L kinetin, 2 mg/L 6-BA and 1 mg/L IAA.     

籼型光敏雄性不育水稻杂种花粉植株游离小孢子培养成植株

禾本科植物游离小孢子的培养已在水稻、小麦、玉米、大麦等主要农作物上获得成功,且在大麦、玉米上成功地从未经预处理及预培养的游离小孢子培养获得了再生植株。籼稻花药培养能力远远低于粳稻,对其游离小孢子的离体培养研究甚少。本文简要报道这方面的研究结果。     

新疆雪莲体细胞胚胎发生

通过体细胞胚胎发生途径实现了新疆雪莲（Saussurea involucrata Kar．et Kir．）的植株再生。选用新疆雪莲子叶为外植体,接种于MS＋0．5mg·L^-12,4-D＋0．05—1mg·L^-1BA的固体培养基上,进行愈伤组织的诱导。从第1次继代培养的愈伤组织中挑选出黄绿色、颗粒状、质地致密的腔陛愈伤组织,转移到含0．05—0．1mg·L^-1 2,4-D的MS液体培养基中进行悬浮培养,20天后可分化产生大量球形胚。继代过程中相继加入PEG和GA3,可以促进体细胞胚的分化和生长。体细胞胚在含有5mg·L^-1 GA3的MS固体培养基上,可发育成完整的植株。     

桔梗的组织培养

桔梗 (ＰｌａｔｙｃｏｄｏｎｇｒａｎｄｉｆｌｏｒｕｍＡ .ＤＣ .)为桔梗科多年生草本植物 ,根供药用 ,有宣肺、祛痰、排脓之功效。用于治疗外感咳嗽 ,咽喉肿痛 ,肺痈吐脓等症[1] 。近年来 ,桔梗成为极具开发前景的一种药食两用经济作物 ,需求量大增 ,野生资源不能满足需要 ,人工栽培问题已有研究[2 ] ,进展也较快。桔梗药材生产主要采用种子或无性繁殖 ,而通过组织培养的方法进行桔梗快繁 ,在 80年代中期少数学者进行过初步研究[3 ,4 ] 。作者拟通过桔梗的离体培养试验 ,优化培养条件 ,为缩短桔梗育苗周期和优良品种快繁提供依据。1　材料…     

Plant regeneration from mesophyll protoplasts ofDianthus superbus

Leaf mesophyll protoplasts ofDianthus superbus were cultured at a density of 5 × 10⁴ protoplasts/ml and divided at about 18% plating efficiency in MS liquid medium supplemented with 0.5 mg/L BAP, 2.0 mg/L NAA and 9% mannitol after 2 weeks. Protocolonies formed after 3 to 4 weeks of culture in the dark at 27°C. These colonies were transferred to continuous illumination (21.5 E m^–2 sec^–1) for 2 weeks where most of the colonies divided to form microcalli, about 2 mm in diameter. Subsequently, green microcalli were transferred to MS solidified medium with 2.0 mg/L 2,4-D that induced shoot-forming calli after 4 weeks. These calli were transferred onto N₆-2 medium containing 0.1 mg/L 2,4-D, 0.1 mg/L NAA, 2.0 mg/L kinetin and 2.0 g/L casein hydrolysate and were cultured under light. After 5 weeks the calli gave rise to multiple shoots (10 to 15 per callus). Upon transfer to MS medium containing 2.0 mg/L NAA, individual shoots were rooted in 4 weeks. The regenerants were successfully transplanted into potting soil.Abbreviations MS Murashige and Skoog - BAP 6-benzylaminopurine - 2,4-D 2,4-dichlorophenoxyacetic acid - NAA 1-naphthaleneacetic acid - N₆ Chu basal salt mixture - MES 2-N-morpholinoethanesulfonic acid     

澳洲青苹组织培养再生体系的研究

通过对澳洲青苹茎尖培养及植株再生体系中无菌培养物的建立、初代培养、继代培养、生根培养和移栽五个技术环节的研究,建立了澳洲青苹组织培养快速繁殖技术体系。结果表明：(1)BA0．3～0．6mg／L NAA0．1～0．3mg／L的MS培养基能促进澳洲青苹茎尖生长或大量侧芽的分化,不定芽生长健壮;(2)无根幼苗在含有IBA0．6～1．0mg／L NAA0．1mg／L的1／2MS培养基上暗培养一周后,再进行自然光培养,30d生根率可达到78%以上;(3)生根苗经50mg／L NAA浸渍1～2h后,移入蛭石：腐殖土：田园土=2：1：1的移栽基质中,保温保湿,成活率达80％以上;(4)进一步建立了自然光下生根、练苗一次完成的同步化体系,缩短了繁育时间,简化了步骤,降低了成本。此技术体系为澳洲青苹优良种苗快速繁殖提供了一条新途径。