盾叶薯蓣组织培养技术的优化

以盾叶薯蓣的根状茎、茎段、叶柄、幼叶为材料,进行愈伤组织诱导、分化及再生植株形成的研究。结果表明：盾叶薯蓣不同外植体均能诱导出愈伤组织,其中茎段愈伤组织的诱导率最高;不同激素配比的培养基对愈伤组织的形成有很大的影响：以LS为基本培养基,2,4-D浓度为4．0mg／L、6-BA浓度为1．0mg／L的激素配比诱导率最高,达62．5％;以改良MS为基本培养基,2,4-D浓度为2．0mg／L、6-BA浓度为0．5mg／L的激素配比诱导率最高,达71．4％。筛选到优化的分化培养基为改良MS附加2．0mg／L的6-BA和0．5mg／L的Vc,且能直接诱导出根,并形成完整植株。     

蔓花生组织培养和植株再生的条件优化

应用MS为基本培养基,通过各种培养条件和不同激素配比,探讨蔓花生组织培养及其植株再生条件的优化。结果显示:幼叶为最佳的外植体,在幼叶愈伤诱导过程中,不超过12h光照,光照强度在21.6μmol.m-2.s-1均可;诱导愈伤组织的适宜培养基为MS+0.5mg/L6-BA+0.2mg/L2,4-D或MS+0.5mg/L6-BA+2mg/LNAA;最适的分化培养基为MS+1mg/LTDZ+2mg/L6-BA+0.5mg/LNAA;最适的生根培养基1/2MS+1mg/LNAA+1mg/LPP333。     

甘蓝型油菜子叶和下胚轴再生植株无性系建立

以甘蓝型油菜(Brassica napus L.)豫油2号和6257的子叶和下胚轴为材料,在不同激素配比的MS培养基上诱导出了愈伤组织。将经过继代的部分愈伤转入分化培养基,分化结果表明:除基因型、外植体和分化培养基的激素配比对分化率有影响外,诱导愈伤培养基的激素配比对分化率也至关重要。豫油2号的子叶和下胚轴在最适诱导培养基(ZT 1+NAA0.5+2,4-D 0.2 mg/L)和最适分化培养基(ZT4+IAA 0.2 mg/L)组合中的愈伤分化率分别为12.5%和75%;6257的子叶和下胚轴在其最适诱导培养基(KT 2+NAA1+2,4-D 0.2 mg/L)和最适分化培养基(6-BA 4+IAA 0.02 mg/L)组合中的愈伤分化率分别为50%和37.5%。将其最适诱导培养基中的愈伤组织继代达8个月以上,建立了不同继代愈伤的再生植株无性系。     

射干真叶、花蕾诱导愈伤组织的研究

目的:建立射干愈伤组织的诱导体系。方法:采用MS培养基为基本培养基,研究不同浓度激素配比及不同AgNO_3浓度对射干真叶、花蕾诱导愈伤组织的影响,并利用显微技术鉴定其愈伤组织类型。结果:射干无菌苗真叶愈伤组织诱导的最佳培养基为:MS+2,4-D 1. 0 mg·L~(-1)+6-BA 1. 5 mg·L~(-1)+NAA 1. 0mg·L~(-1)+AgNO_315mg·L~(-1),花蕾愈伤组织诱导的最佳培养基为:MS+2,4-D 1. 0 mg·L~(-1)+6-BA 1. 0 mg·L~(-1)+NAA 1. 0 mg·L~(-1),诱导率均高于60%,其中2,4-D对射干愈伤组织诱导至关重要,AgNO_3对射干真叶胚性愈伤组织的形成有一定的影响。结论:花蕾是射干愈伤组织诱导较理想的外植体材料,获得的高效愈伤组织诱导条件将为后续射干细胞悬浮体系的成功构建奠定基础。     

鱼腥草体细胞胚胎发生和植株再生

目的:利用鱼腥草的叶片和叶柄为材料,进行体细胞胚胎诱导及植株再生研究。方法:运用正交设计试验,考察在改良的MS固体培养基上添加不同种类、不同浓度的植物生长物质组合及其配比对鱼腥草愈伤组织诱导、体细胞胚胎发生及植株再生的影响。结果:鱼腥草无菌苗叶片在含有2,4-D 1.0 mg/L 6-BA 0.5 mg/L的改良MS培养基上能诱导出胚性愈伤组织;胚性愈伤组织在含有6-BA 1.0 mg/L的改良的MS培养基上诱导体细胞胚的发生;叶柄在含有6-BA 1.0 mg/L改良MS培养基上直接产生体细胞胚。体细胞胚在改良的MS NAA0.1 mg/L 6-BA 1.0 mg/L的培养基上能够快速繁殖,形成大量不定芽,在不加任何激素的MS培养基上就可以萌发出不定根,发育为成完整植株,在MS IBA 1.0 mg/L的固体培养基上能够形成大量的根。结论:建立了鱼腥草体细胞胚胎发生及植株再生的体系。     

冬凌草离体培养体系的建立及主要次生代谢产物的测定

以冬凌草叶片为外植体,研究不同浓度激素组合对冬凌草愈伤组织诱导及植株再生的影响,并对不同外植体(茎、叶)诱导愈伤、芽的分化能力及再生植株内主要次生代谢产物的含量进行了比较研究。结果表明:在MS 2.0 mg/L 6-BA 1.0 mg/L NAA培养基上诱导愈伤组织效果较好;在MS 2.0 mg/L 6-BA的培养基上诱导芽的效果较好;叶片和茎段在愈伤诱导培养基上均能产生大量的愈伤组织,但其再分化能力以茎段最好;再生苗生根培养基以0.3 mg/L IBA最好;以叶为外植体诱导的再生植株中冬凌草甲素、迷迭香酸的含量均高于以茎为外植体诱导的再生植株。     

百合体细胞胚胎发生和植株再生

以切花百合(Lilium)品种‘黄天霸’(‘Manissa’)花器官为外植体诱导体细胞胚胎发生与植株再生。结果表明,不同花器官、不同激素配比对愈伤组织形成均具有显著影响。花丝为最佳外植体,激素对愈伤组织诱导的影响效应为NAA>6-BA>2,4-D,最适培养基为MS+1.0 mg.L-1NAA+0.2 mg.L-16-BA;激素诱导体细胞胚胎发生的影响效应为2,4-D>KT>6-BA,最佳培养基配方为MS+1.0 mg.L-12,4-D+0.2 mg.L-1KT+1.0 mg.L-16-BA;MS培养基添加IBA可促进体细胞胚萌发成苗,体细胞胚芽成苗的最佳培养基为MS+0.2 mg.L-16-BA+1.0 mg.L-1IBA。     

曼陀罗茎段愈伤组织诱导和再生植株的研究

本试验以曼陀罗茎段为外植体,在附加不同植物激素组合的培养基中对愈伤组织的诱导和植株再生进行研究。结果表明:采用修改的MS培养基(除去甘氨酸,维生素B1含量增加至0.5mg/L,pH5.5)附加2mg/L2,4-D可由曼陀罗茎段诱导大量胚性愈伤组织;愈伤组织继代选用0.5mg/L2,4-D为宜;不定芽的诱导采用MS培养基(20g蔗糖,8g琼脂,0.1g水解干酪素) 6-BA(0.5mg/L);幼苗进一步转接至1/2MS IBA(0.2mg/L)生根培养基中,可完成曼陀罗茎段愈伤组织诱导和再生植株的组织培养过程。     

轮叶党参的组织培养及植株再生研究

以轮叶党参为材料,研究了不同外植体、激素组合、培养基及光照条件对愈伤组织诱导及植株再生的影响.结果表明,叶片是轮叶党参组织培养较为合适的外植体.外植体在添加0.5 mg·L~(-1) 2,4-D+1.0 mg·L~(-1) 6-BA+0.5 mg·L~(-1) KT的MS培养基上愈伤组织诱导率最高可达100%;愈伤组织转移到附加0.75 mg·L~(-1) 6-BA+0.5 mg·L~(-1) NAA的MS培养基进行继代培养,增殖后的愈伤组织转移到附加0.5 mg·L~(-1) 6-BA+0.2 mg·L~(-1) NAA的1/2MS分化培养基进行分化,其分化率达89.4%;将分化出的芽转接到附加0.2 mg·L~(-1) IAA+0.2 mg·L~(-1) NAA的1/2MS培养基,生根率达92.5%.暗培养诱导出愈伤组织后转到16 h·d~(-1)光照条件下,愈伤组织增殖倍数和分化率显著提高,再生苗健壮,长势强.     

党参的离体培养及植株再生的研究

在附加激素的MS培养基上,培养党参下胚轴和无菌芽切段,诱导产生愈伤组织并且再生植株。经过两年多(15个世代)的继代培养,建立了党参体细胞无性系。实验结果表明:(1)培养基MS 0.4mg/L 2,4-D 0.8mg/L Kt 2.0mg/L IAA对愈伤组织诱导及继代培养,MS 0.2mg/L 6-BA诱导外植体产生丛芽和愈伤组织再分化,MS 0.5mg/L NAA 0.2mg/L 6-BA及MS 0.2mg/L NAA诱导生根效果最好。(2)愈伤组织再分化经过胚状体途径。     

在1/3海水培养基上筛选豆瓣菜耐盐变异体

The responses of stem segments of watercress ( Nasturtium offtcinale R. Br. ) to 6-BA, NAA and 2,4-D were studied. MS medium supplemented with 2.0 mg/L 6-BA, 0.2 mg/L 2,4-D was used for callus initiation and maintainance. MS medium supplemented with 4.0 mg/L 6-BA was suitable for plant regeneration and MS medium without plant hormone supplement was used for rooting and plant propagation. For screening of salt. tolerant calli, stem segments of watercress were plated onto callus initiation medium containing 1/3 natural seawater. Seventeen out of the 325 plated explants produced calli. The growth curves demonstrated that the growth rate of salt-tolerant calli on saline medium almost matched that of the control calli on normal medium. Some of the salt-tolerant calli were transferred to the normal regeneration medium or saline regeneration medium to induce plant regeneration. In the first case, buds and shoots were regenerated in the same way as those of control calli on normal regeneration medium. More than 1 000 regenerated shoots were obtained of which 83 regenerated shoots were cut and transferred to saline MS base medium. At first, all shoot growth was inhibited, but 40 days after the transfer, rapid-growing axillary shoots were observed on 16 of the original shoots but none on the control shoots on saline MS base medium. Moreover, green spots appeared on most calli 10 days after they were transferred to saline medium, however buds appeared only on 5 calli from the 30 transferred calli and at the end only 2 rapid-growing shoots were obtained from two calli. In total, 18 variant lines were obtained through propagation of the salt-tolerant shoots on saline MS base medium. RAPD analysis was performed in 10 of the 18 salt-tolerant variant lines and DNA variation was detected in all the tested variant lines.     

二棱大麦体细胞胚性无性系的建立

对１２个品种的二棱大麦（Ｈｏｒｄｅｕｍｄｉｓｔｉｃｈｕｍ２ｎ＝２４）幼穗和６个品种的幼胚在附加５００ｍｇ／Ｌ酪蛋白,１５０ｍｇ／Ｌ天门冬素和２ｍｇ／Ｌ２,４－Ｄ的ＭＳ培养基（Ｍ＿（３２０））中进行了离体培养,并诱导产生出了愈伤组织。把由Ｄ＿１幼胚所获得的愈伤组织转移到含２,４－Ｄ０．５ｍｇ／Ｌ（Ｍ＿（３０５））培养基中后诱导产生出了体细胞胚性愈伤组织,幼苗和根。筛选出了体细胞胚性无性系。在长达二年的继代培养中,该细胞系仍保持旺盛的再生和分化成苗能力。     

荞麦组织培养及高频植株再生体系的建立

通过对荞麦(Fagopyrum esculentum Moench)不同外植体、不同激素配比的比较研究,建立了荞麦离体培养高效植株再生体系。荞麦子叶切段在含2．0 mg/L 2,4-D和1．0 mg/L 6-BA的MS培养基上愈伤组织诱导率为89．6％,而下胚轴切段在含2．0 mg/L 2,4-D和1．0-2．0 mg/L 6-BA MS培养基上愈伤组织诱导率高达100％。在2．0 mg／L 6-BA、0．1 mg,L IAA和1 mg／L KT的MS培养基上通过愈伤组织间接分化或外植体直接分化形成不定芽。来自子叶和下胚轴的愈伤组织的分化率分别为42．5％和73．6％,下胚轴的分化率明显高于子叶。将生长状态良好的不定芽转至含1．0 mg/L IBA和0．5mg/L NAA的1/2 MS培养基上生根,生根率达到100％。再生植株移栽到盆土中,成活率达91．6％,并且生长状态和特征均表现正常。     

Direct and indirect regeneration of plants from internodal and leaf expiants of<Emphasis Type="Italic">Hypencum bupleuroides</Emphasis> gris

Species of the genusHypencum are of considerable interest worldwide because of their medicinal properties.In- vitro culture is a useful tool for both multiplication of the genus and studying its economically important secondary metabolites. Here, we present an effectivein- vitro propagation method forH. bupleuroides. Leaf and internodal expiants excised from 9-week-old,in vitro-germinated seedlings were cultured on a Murashige and Skoog (MS) medium supplemented with benzyladenine (BA; 1.0 or 0.1 mg L^-1) and 2,4-dichlorophenoxyacetic acid (2,4-D; 1.0 or 0.1 mg L¹). Depending on the BA and 2,4-D combination used, these cultures produced adventitious shoot buds directly on the surfaces of both types of explants as well as excessive calli. Numerous shoots were obtained when the calli from both expiant types were cultured on an MS medium supplemented with 2 mg L^-1 BA. Internodal expiants were more responsive than leaf tissues to direct and indirect plant regeneration. After shoots that regenerated from either the calli or the expiant surface were excised, rooting was best on an MS medium lacking any growth hormones. These rooted plants were then acclimatized under greenhouse conditions, and 90% of regenerants had survived. Ours is the first report ofin- vitro plant regeneration fromH. bupleuroides.     

欧美黑杨离体再生途径及影响因子的研究

先期完成了以腋芽发育为再生方式的研究后 [1 ] ,又对不定芽的发育进行了探索 .分别以欧美黑杨特选品系的不同部位外植体进行愈伤组织诱导及植株再生研究 ,易诱导并易分化出新芽的外植体为幼嫩茎段 ;通过不同激素浓度合理配比 ,进行适宜的诱导及分化培养基筛选 ,培养基为 MS+6 - BA 1.2 mg/ L +NAA 0 .5 mg/ l,附加 6g/ L琼脂 ,诱导的愈伤组织分化出密集的再生芽 ;糖分在诱导分化过程中作用突出 ,其合适浓度为 4 0 g/ L ;愈伤组织的形态及培养时间对分化频率影响较大 ,最佳的形态为致密的绿色 ,白色及粉红色的愈伤组织较疏松 ,分化率很低 .不同部位外植体、激素含量、蔗糖浓度、愈伤组织形态及培养时间是明显影响欧美黑杨愈伤组织诱导分化的因子 .     

Agrobacterium rhizogenes-mediated transformation and the regeneration of transformants in Alhagi pseudalhagi]

The regenerated shoot segments of Alhagi pseudalhagi were sliced and infected with Agrobacterium rhizogenes strain A4. The hairy roots and transformed calli were obtained through selection on hormone free MS medium. The transformants were cultured on MS medium with 2 mg/L 2,4-dichlorophenoxy acetic acid (2,4-D) and 0.5-1 mg/L 6-benzylaminopurine (6-BA) to induce calli. 3 mg/L 6-BA and 0.5 mg/L naphthalene acetic acid (NAA) were applied for shoot differentiation. Shoots were planted on MS medium with 2 mg/L indole-3-butyric acid (IBA) and produced roots. Opine analysis proved the integration and expression of T-DNA in over 95% hairy roots, 75% transformed calli and transformed plantlets respectively. The 81% hairy root cells had normal chromosome numbers (2n = 18). The alterations of chromosome number were observed. After one year of subculturing, the regeneration ability of transformants was maintained.     

Plant regeneration from protoplasts of hydroxyproline resistant cell line in Onobrychis viciaefolia

An efficient protocol for plant regeneration from protoplasts of hydroxyproline(HYP)resistant cell line of Onobrychis viciaefolia was established.In SH medium supplemented with 1mg/L2,4-dichlorophenoxy-acetic acid(2,4-D),0.5mg/L kinetin(KT)and 0.2mg/L naphthalene acetic acid(NAA),the division frequency of protoplastderived cells reached up to over 60%,and microcalli were obtained in 5-6wk.Upon transferring them on agar solidified MS medium plus 2mg/L indole-3-acetic acid (IAA),shoots were induced.After cultivating them on MS medium with or without IAA,roots were regenerated.Chromosome number of all protoplast-regenerated plants examined were normal(2n=28).The protoplast-derived calli and plants grew vigorously on the medium containing 10 mmol/L HYP.     

枸杞原生质体培养及高效成株体系的建立

由枸杞髓部组织诱导出胚性愈伤组织,并由此愈伤组织建立起稳定的细胞悬浮系。从悬浮细胞游离的原生质体在改良KM培养基(1.5 mg/L 6_BA,0.5 mg/L NAA和0.5 mg/L 2,4_D)中进行液体浅层培养,3～4 d后出现第一次分裂,第7 d统计分裂频率为50.3%,15 d左右可形成细胞团,3～4周后形成肉眼可见的愈伤组织,愈伤组织植板率为1.25%。将细胞团转移到液体分化培养基(MS+6_BA 1.5 mg/L+2,4_D 0.2 mg/L) 8～10 d可形成大量胚状体,及时将胚性愈伤组织块转移到固体分化培养基上(MS+6_BA 0.2 mg/L),可形成大量绿芽,分化率54.17%。绿芽在生根培养基（MS+NAA 0.2 mg/L）可形成完整植株,移栽后成活良好。     

Factors influencing induction,propagation and regeneration of mature zygotic embryo-derived callus from Allium cepa

A systematic study on the effects of subspecies, cultivar, basal medium, sucrose concentration and 2,4-dichlorophenoxyacetic acid concentration on callus induction, propagation and subsequent plant regeneration in Allium cepa has been carried out. Mature zygotic embryos from two onion (cvs. Sturon and Hyton) and two shallot (cvs. Tropix and Atlas) varieties were used as explants. After callus initiation and growth on both Murashige and Skoog (MS) and Gamborg's B5 modified by Dunstan and Short (BDS) basal media with different 2,4-dichlorophenoxyacetic acid and sucrose concentrations for eight weeks, lines were identified on which compact or friable callus was induced. Callus induction and propagation were largely determined by the concentration of 2,4-dichlorophenoxyacetic acid whereas subspecies, cultivar, sucrose concentration and basal media were of less importance. After callus propagation for twelve weeks, 315 lines from a total of 3348 embryos initially subcultured were selected to test their regeneration capacity on growth regulator-free medium. It was found that shallot formed more shoots and roots than onion. The MS basal medium proved to be more beneficial for shoot regeneration and root formation than the BDS basal medium. There were no differences in plant regeneration among selected calli which had been previously subcultured on different concentrations of 2,4-dichlorophenoxyacetic acid and sucrose. The results show that plant regeneration strongly depended on the line: 45.4% from 315 tested lines could produce shoots while 93.0% formed roots. This revised version was published online in June 2006 with corrections to the Cover Date.