半海水筛选耐盐紫花苜蓿细胞系的生理特性分析

Physiological Characteristics Analysis of Salt-tolerant Cell Line from Alfalfa Callus Cultures Selected on Half Natural Seawater Medium

以紫花苜蓿无菌苗下胚轴为外植体,诱导获得愈伤组织(M0)。经甲基磺酸乙酯(EMS)处理,通过一步筛选获得了耐半海水w(NaCl≈1.5%,1/2海水)的耐盐细胞系(Ms1),一部分耐盐细胞系转移到无盐培养基上,继代培养10代后得到新细胞系(Ms2)。无论是在无盐还是不同程度盐胁迫条件下,Ms1的成活率、生长率、脯氨酸含量、Cl-、K+、K+/Na+、可溶性蛋白质含量都高于M0,差异显著。Ms2介于两者之间,但是与Ms1基本接近,差异不显著。在w(NaCl≈0.6%,1/5海水)条件,M0细胞系的SOD活性高于Ms1和Ms2;在w(NaCl≈1%,1/3海水)条件下,Ms1、Ms2细胞系的SOD活性达到最高,与M0可溶性蛋白质SDS-PAGE图谱相比较,Ms1、Ms2中出现两条新多肽带(P1=58kD,P2=26kD),这两条多肽带可能与耐盐性有关。

M0 cell line of alfalfa （Medicago sativa L. ） was induced from alfalfa aseptic hyopocotyl segments. Salt-tolerant cell line （Msl） of alfalfa to w（NaCl≈1. 5%, half seawater） was obtained directly after dealing with EMS. After some Ms1 was subcultured through 25 passages in a medium containing half natural seawater, then transferred to NaC1 free medium for 10 passages and the Ms2 cell line was gained. The results of physiological characteristics analysis showed, whether normal condition or salt stress, the survival rate of calli （SRC）, growth rate of calli （GRC）, the contents of proline, K^＋ , and Clˉ, the rate of Ms2 and Ms2 were significantly higher than those of M0. In despite of above indexes of Ms2 were between M0 and Ms1, but all were near to Ms1 , and there was no remarkable difference with Ms1. In normal condition, the SOD activity and Na＋ of M0 were lower than Ms, and Ms2. In condition of lower seawater concentration, the SOD activity and Na＋ of M0 were higher than Msl and Ms2. SDS-PAGE electrophoresis of soluble proteins indicated that there were two new protein bands in Ms1 and Ms2 compared to the control. These proteins were 58kD and 26kD and they might be related to salt tolerant.