西洋参细胞大量培养的研究

当西洋参细胞培养在Ms培养液中,KN0₃含量提高一倍而去掉NH₄NO₃。时细胞生长速率和皂甙产量分别比在正常培养液中提高65．1％和166．2％。黑节草寡糖素和人参寡糖素均有利于西洋参细胞的生长和皂甙含量的提高,尤其能增加Rg组皂甙的含量。西洋参细胞悬浮培养以生产皂甙收获的最佳时期为培养25天以上,其合成皂甙的高峰在细胞生长的对数期稍后出现。细胞悬浮培养和发酵培养过程中均未见pH值回升的现象。pH值稳定的发酵培养和pH值任其变化的培养相比,其皂甙含量,生长速率和生物量均要高。最后对细胞的培养方式进行了比较。     

三七细胞大量（发酵）培养的研究

三七(Panax notoginseng)细胞发酵培养的皂甙含量为干重的11.21%,皂甙产率为1513.3mg·L~(-1),细胞培养物产率为每月13.58g干重·L~(-1),均比悬浮培养高。接种量的增加能明显地提高生长速率、皂甙产率以及细胞培养物产率。通气速率0.8vvm较适合于三七细胞的发酵培养。在发酵培养过程中pH值由5.8逐渐降至3.92,没有出现回升。     

三七.人参和西洋参细胞悬浮培养的比较研究

用薄层层析对三七、人参和西洋参愈伤组织进行的初步鉴定表明,三种愈伤组织都含有皂甙和主要皂甙成分Rb_1、Rg_1,三七愈伤组织还含有一种抗癌皂甙Rh_1。对愈伤组织的生长,三七低于人参高于西洋参;对愈伤组织中总皂甙含量,三七均高于人参和西洋参。三种植物细胞悬浮培养结果类似于他们的愈伤组织培养,但生长又进一步提高。三七细胞悬浮培养中皂甙产生的时间进程几乎与生长平行,合适的收获期为培养30天。寡糖素不仅增强三七培养细胞的皂甙形成而且促进细胞生长,较合适的浓度为1.25 ppm。通过以上研究,使三七悬浮培养细胞的生长(干重增加178毫克)为最初培养愈伤组织的4倍以上,总皂甙产率高达20.6毫克,为最初培养愈伤组织的8.5倍。     

西洋参细胞液体培养及动力学研究

文对西洋参细胞在改良MS培养液中发酵过程和发酵动力学进行了研究。结果表明,细胞生长动力学为：dXdt=μmax·s Ksx,式中μmax=o．1613d^-1,Ks=47．59g／L,基质消耗动力学为。-dsdt=1．75dXdt。西洋参细胞液体培养收获最佳期23天以上,皂甙累积的高峰期为25天以后。液体培养时,细胞中大部分皂甙释放到培养液中,经薄层层析后仅含R_g1;而愈伤组织中则含Rg₁、Rb₁、Re、Rd、Re五种,其总皂甙含量比人工栽种西洋参提高23％。     

西洋参悬浮细胞发酵工艺研究

探讨了西洋参悬浮细胞分步培养与稀土、D-半乳糖和甘露醇等诱导子对悬浮细胞生长及皂甙产量的影响。发现继代4d后换液一次再继续培养获得的培养物,在皂甙产率和糖利用率等方面优于连续培养;D-半乳糖作为诱导子,对悬浮培养的西洋参细胞生长、皂甙产率及皂甙的分泌等方面都有非常明显的促进作用。     

西洋参细胞悬浮培养中皂甙生物合成的代谢调节

七种真菌菌丝体诱导因子中,六种促进培养细胞的皂甙生物合成。尤其葡枝根霉作用更为明显,提高皂甙含量二倍。人参寡糖素是一种既能诱导皂甙生物合成,又能促进细胞生长的新型诱导因子,浓度为50mg／L寡糖素既能维持细胞较好生长又能提高皂甙含量,因而得到较高的皂甙产率。用四种条件培养液进行培养,结果均促进悬浮培养细胞的皂甙生物合成,其中丹参条件培养液使悬浮培养细胞的皂甙含量提高二倍多。加入皂甙生物合成主途径的三种前体,都不同程度地促进皂甙的生物合成,前体中以角鲨烯作用最为明显。     

云南红豆杉细胞发酵培养的研究

利用云南红豆杉(Taxus yunnanensis)细胞悬浮培养的最佳培养条件,进行了细胞10L规模的发酵培养研究。对细胞发酵培养过程中的pH值变化、糖利用率以及细胞生长周期等实验参数进行了测定。已进行的3次发酵培养的实验结果表明:培养细胞的生长率已达0.4g/L.d(即12.0g/L),培养细胞中紫杉酵的含量为0.119％(其中培养液中的紫杉醇含量约占42％)。初步建立了优化的云南红豆杉细胞大量培养系统。     

真菌激发子对人参悬浮培养细胞的生长和人参皂甙生物合成的影响

真菌诱导子处理人参悬浮培养细胞后,人参皂甙的合成有明显增加,诱导处理改变人参皂甙的积累时程,促进人参细胞培养物中次生产物的外泌,同时增强细胞对蔗糖的摄取、吸收并引起细胞H~ 流的变化。     

西洋参细胞大量培养的工艺学研究

较适合于植物细胞悬浮培养的培养液体积为三角瓶总容量的1／5至2／5。氧溶浓度的输出随着温度的增加而增加。西洋参细胞培养液的粘性明显大于无细胞培养液。细胞发酵培养较合适的搅拌速度为60rpm。渗透压的增大会明显地提高细胞培养物的皂甙含量,但对细胞的生长不利。本研究为今后设计出适合于西洋参细胞大量培养的发酵罐提供了重要的依据。     

B群链球菌培养条件的优化研究

利用B群链球菌(Streptococcus Group B,SGB)有可能发展一种新的预防用疫苗。为研究其生长特性,对该菌在不同培养条件下的生长状况进行了研究。分别考察了液体培养基、接种量、pH值、生长因子、溶氧等因素对SGB生长的影响。结果发现：SGB液体培养基生长因子中尿嘧啶、烟酸、泛酸钙等对SGB的生长有较大影响,葡萄糖最佳浓度为14g/L;此外,发酵培养最佳接种量为0．5～0．8亿／ml;最适pH值为7～8,培养过程中调节pH、补加葡萄糖能维持SGB继续生长;增大溶氧对SGB生长影响不明显。在此优化基础上,连续进行3批100L发酵,SGB菌生长良好,荚膜多糖产量可观。     

Embryo rescue inVicia faba andVicia narbonensis

Embryo rescue in twoVicia faba L. cultivars (‘Polycarpe’ and ‘A-107’) and oneV. narbonensis L. population (A-202) was studied under a 22 ± 2°C/ 16 ± 1° day/night temperature regime. Very young ovules (1.0–1.8 mm long) cultured, in-ovule, on five liquid media remained green for a longer period of time on modified B5, modified Murashige and Skoog and modified Beasley and Ting media than on modified Phillips and Collins and modified Bourgin and Nitsch media. However, no embryo growth or embryo germination was observed. In-ovule culture of older ovules, 6 and 8 days forV. narbonensis and 10 and 14 days forV. faba, on modified B5 liquid medium allowed 6-day-oldV. narbonensis and 14-day-oldV. faba embryos to be rescued. Finally, culture of whole pods of the two species resulted in the rescue of even younger embryos. Thus, plantlets were obtained from as young as 4-day-oldV. narbonensis pods and 11-day-oldV. faba pods.     

二十世纪我国植物学家对植物组织培养的贡献

回顾了上一世纪我国植物组织培养的发展.1934年以来,我国的植物组织培养研究一直与国际发展同步进行.我国学者在离体器官发生、茎尖培养、花药培养、子房培养、胚乳培养、原生质体培养和细胞大量培养等分支领域都取得重要进展.本文在引证我国研究者发表的植物组织培养论文的基础上,着重评述了那些被国际同行公认的研究成果.此外,还介绍了植物组织培养在我国农业和工业上应用的情况.     

二十世纪我国植物学家对植物组织培养的贡献

回顾了上一世纪我国植物组织培养的发展。 1934年以来 ,我国的植物组织培养研究一直与国际发展同步进行。我国学者在离体器官发生、茎尖培养、花药培养、子房培养、胚乳培养、原生质体培养和细胞大量培养等分支领域都取得重要进展。本文在引证我国研究者发表的植物组织培养论文的基础上 ,着重评述了那些被国际同行公认的研究成果。此外 ,还介绍了植物组织培养在我国农业和工业上应用的情况     

猪苓、伴生菌及蜜环菌共培养的形态学研究

本文对猪苓、伴生菌及蜜环菌两两共培养及三者共培养进行了宏观形态观察及细胞学水平上的研究。结果表明,猪苓与伴生菌共培养时,在二者之间形成一致密拮抗线,猪苓菌落表面菌丝分化产生大量菌丝束;猪苓与蜜环菌共培养时,猪苓能阻止蜜环菌菌索对其自身的进一步侵袭,互作区中的双方菌丝及菌索均停止生长;蜜环菌与伴生菌共培养时,蜜环菌能穿透整个伴生菌菌落,在伴生菌菌落下方产生大量分枝;三者共培养后,猪苓对蜜环菌的防御能力有所下降,伴生菌对蜜环菌的耐受力有所提高,蜜环菌产生的新分枝均向伴生菌一侧生长,猪苓与伴生菌之间并不形成致密拮抗线,只可见双方菌丝的白色交融区。 猪苓与伴生菌均能在蜜环菌菌索皮层上形成侵入位点。     

Tapping culture-an improved method for cell suspension culture

Summary A new culture vessel was designed for cell suspension culture. A silicone-convered magnet bar fixed by one end to the side wall of the bottle was held horizontally a short distance from the bottom. A standard type magnetic stirrer was used. In contrast to the conventional horizontal movement of “stirring” in cultures the bar moves vertically with a “tapping” motion. This improvement resulted in less cell injury, higher rate of cell proliferation and formation of fewer bubbles than in the conventional type. Nine cell types were simultaneously cultivated in tapping, stirring and stationary culture. All cell types proliferated more luxuriously in tapping cultures than in stirring cultures. Serial cultivation of cells in tapping cultures was also successful. This work was supported in part by the grants for Cancer Research from the Ministry of Education, Science and Culture, Japan.     

Light requirement and photosynthetic cell cultivation – Development of processes for efficient light utilization in photobioreactors

Although the potential of photosyntheticmicroorganisms for production of various metabolitesand in environmental bioremediation is recognized,their practical application has been limited by thedifficulty in supplying light efficiently tophotobioreactors. Various types of photobioreactorwith high illumination to volume ratios have beenproposed, but most are limited by cost, mass transfer,contamination, scale-up or a combination of these.The problem of light supply to photobioreactorscan be solved by developing photosynthetic cellcultivation systems where light is either substitutedor supplemented. Many strains of photosynthetic cellsare capable of heterotrophic growth under darkconditions and their heterotrophic culture can be usedfor efficient production of biomass and somemetabolites. However, light is absolutely required forefficient production of some metabolites. In suchcases, there is a need to supplement the heterotrophicwith photoautotrophic metabolism. Inphotoheterotrophic (mixotrophic) culture, thephotoautotrophic and heterotrophic metabolisms can beexploited for efficient production of usefulmetabolites but it has many problems such as processoptimization in terms of making a balance between thephotoautotrophic and heterotrophic metabolism. Another promising system is the sequentialheterotrophic/ photoautotrophic cultivation system,where the cells are cultivated heterotrophically tohigh concentrations and then passed through aphotobioreactor for accumulation of the desiredmetabolite(s). Furthermore, cyclicphotoautotrophic/heterotrophic cultivation system canbe used to achieve continuous cell growth underday/night cycles. This involves cultivating thecells photoautotrophically using solar light duringthe day and then adding controlled amount of organiccarbon source during the night for heterotrophicgrowth. In this review, these various systems arediscussed with some specific examples.     

荔枝生物技术研究进展（综述）

从荔枝的组织培养、花药培养和原生质体培养方面综述荔枝生物技术的研究概况,并提出该领域存在问题及发展方向。     

Technology for cell cycle research with unstressed steady-state cultures

A culture system for performing cell cycle analyses on cells in undisturbed steady-state populations was designed and tested. In this system, newborn cells are shed continuously from an immobilized, perfused culture rotating about the horizontal axis. As a result of this arrangement, the number of newborn cells released into the effluent medium each generation is identical to the number of cells residing in the immobilized population, indicating that one of the two new daughter cells is shed at each cell division. Thus, the immobilized cells constitute a continuous, steady-state culture because the concentrations, locations and microenvironments of the cells in the culture vessel do not vary with time. In tests with mouse L1210 lymphocytic leukemia cells, about 10⁸ newborn cells were produced per day. This new culture system enables a multiplicity of cell cycle analyses on large numbers of cells assured to be from populations in steady-state growth.

A simple device for planting replicate mammalian cell cultures

Summary The design and use of a unit for planting uniform inocula for replicate cultures are described. Its design permits continuous gassing of suspensions of mammalian cells with humidified CO₂, thus stabilizing the pH (±<0.05 pH unit) of culture media buffered with sodium bicarbonate. The unit can be readily modified to deliver different volumes; identical samples can be dispensed simply and rapidly, with minimal cell damage and chance of microbial contamination. Quantitative data regarding sample uniformity and growth subsequent to planting with this unit are presented.     

A simple device for planting replicate mammalian cell cultures

Summary The design and use of a unit for planting uniform inocula for replicate cultures are described. Its design permits continuous gassing of suspensions of mammalian cells with humidified CO₂, thus stabilizing the pH (±<0.05 pH unit) of culture media buffered with sodium bicarbonate. The unit can be readily modified to deliver different volumes; identical samples can be dispensed simply and rapidly, with minimal cell damage and chance of microbial contamination. Quantitative data regarding sample uniformity and growth subsequent to planting with this unit are presented.