Datko A H, Mudd S H, Giovanelli J
National Institute of Mental Health, Bethesda, Maryland 20205.
Plant Physiol. 1980 May;65(5):906-12. doi: 10.1104/pp.65.5.906.
Photoautotrophic and mixotrophic growth of Lemna paucicostata Hegelm. 6746 (formerly Lemna perpusilla Torr. 6746) was investigated to establish standardized conditions for biochemical studies. Optimal temperature for growth was 29 to 30 C. The medium used previously (Datko AH, Mudd SH, Giovanelli J 1977 J Biol Chem 252: 3436-3445) was modified by inclusion of NH(4)Cl, decreasing macronutrient and ethylenediamine tetraacetate concentration, increasing micronutrient concentration, and inclusion of bicarbonate (for photoautotrophic growth) or 2-(N-morpholino)ethanesulfonic acid (for mixotrophic growth) buffers. Varying the sulfate concentration between 14 and 1 millimolar had no effect on growth. For photoautotrophic growth in the new medium (medium 4), the effects of CO(2) concentration, light intensity, and pH were measured. Under the optimal conditions, a multiplication rate (MR) of 300 to 315, equivalent to a doubling time of 23 to 24 hours was obtained. Addition of glutamine or asparagine did not increase this MR. For mixotrophic growth in low light, the effects of sucrose concentration and pH were determined. Under optimal conditions, MR was 210. A concentration of sucrose less than maximal for growth was chosen for the medium for experiments which will include (14)C-labeling of intermediates. MR under these conditions was 184. Growth was equally good in medium 4 and in half-strength Hutner's medium when sulfate was high (0.4 to 1 millimolar), but better in medium 4 when sulfate was low (20 micromolar). Growth rates could be restored to normal in half-strength Hutner's with low sulfate by decreasing the molybdate concentration.By modifying medium 4 to contain very low amounts of sulfate, and by preconditioning medium and plants, it was shown that there was an increment in plant protein of approximately 2.5 micrograms per nanomole of added MgSO(4). Colonies undergoing sulfur limitation exhibited a slow growth rate and a high frond to colony ratio. Molybdate and selenate produced growth inhibition reversible by sulfate. Conditions were developed in which the plants could be maintained indefinitely in the presence of either molybdate or selenate in altered metabolic steady-states with lowered growth rates and protein per frond.
为了建立用于生化研究的标准化条件，对少脉浮萍Lemna paucicostata Hegelm. 6746（以前称为Lemna perpusilla Torr. 6746）的光合自养生长和混合营养生长进行了研究。生长的最佳温度为29至30摄氏度。先前使用的培养基（Datko AH，Mudd SH，Giovanelli J 1977 J Biol Chem 252：3436 - 3445）通过加入氯化铵、降低大量营养素和乙二胺四乙酸浓度、增加微量营养素浓度以及加入碳酸氢盐（用于光合自养生长）或2 -（N - 吗啉代）乙磺酸（用于混合营养生长）缓冲液进行了改良。将硫酸盐浓度在14至1毫摩尔之间变化对生长没有影响。对于在新培养基（培养基4）中的光合自养生长，测量了二氧化碳浓度、光照强度和pH值的影响。在最佳条件下，获得了300至315的增殖率（MR），相当于23至24小时的倍增时间。添加谷氨酰胺或天冬酰胺并没有提高这个MR。对于低光照下的混合营养生长，测定了蔗糖浓度和pH值的影响。在最佳条件下，MR为210。对于将包括中间体的（14）C标记的实验，选择了低于生长最大浓度的蔗糖浓度用于培养基。在这些条件下的MR为184。当硫酸盐含量高（0.4至1毫摩尔）时，在培养基4和半强度Hutner培养基中的生长同样良好，但当硫酸盐含量低（20微摩尔）时，在培养基4中的生长更好。通过降低钼酸盐浓度，可以使低硫酸盐的半强度Hutner培养基中的生长速率恢复正常。通过将培养基4改良为含有极低量的硫酸盐，并对培养基和植物进行预处理，结果表明，每添加1纳摩尔硫酸镁，植物蛋白质增加约2.5微克。经历硫限制的群体表现出缓慢的生长速率和高的叶状体与群体比率。钼酸盐和硒酸盐产生的生长抑制可被硫酸盐逆转。已经开发出一些条件，在这些条件下，植物可以在存在钼酸盐或硒酸盐的情况下无限期维持，处于代谢稳态改变、生长速率降低和每叶状体蛋白质含量降低的状态。
