S180-S2D9传代细胞的DNA指纹图谱、细胞核型、生物学特性及DNA含量的比较研究

The Comparison Study in DNA Finger map,Karyological Characteristics,Biology Characteristics and DNA Contents of S180-S2D9 from Defferent Generations

目的:通过比较S180-S2D9传代细胞的DNA指纹图谱、细胞核型、生物学特性及DNA含量,对其进行质量控制.方法:提取S180-S2D9传代细胞的DNA后,进行RAPD分析;细胞培养加秋水仙碱,涂片、染色后,显微镜下计数染色体数目;原代、50代、100代细胞分别接种KM小鼠腹腔,观察腹水的生长情况及小鼠的存活天数;培养的细胞用70%乙醇固定,流式细胞仪测DNA含量.结果:与原代、50代细胞相比,100代细胞的DNA指纹图谱发生变异;50代、100代、100代克隆株1、100代克隆株2、100代克隆株3,其染色体均数做方差分析表明,两两单位比较均无显著统计学差异(P>0.05).原代、50代、100代细胞分别接种于10只昆明小鼠腹腔,3组的存活天数分别为13～23d、13～20d、10～14d,与原代、50代相比,100代均数有显著统计学差异(P<0.05),原代与50代均数比较无显著统计学差异(P>0.05).流式细胞术DNA含量分析表明,5株细胞的相对分子质量分别为0.3575、0.3984、0.3542、0.3919、0.3890.结论:在细胞核型与DNA含量的比较上,5株细胞无显著统计学差异;在DNA指纹图谱与对小鼠致死性方面提示传代细胞发生了变异.

Objective： By comparing the DNA Finger map, Karyological Characteristics Biology Characteristics and DNA Contents of S180- S2D9 from defferent generations, they are quality - controlled.Methods： To extract DNA and analyze DNA Finger map of $180- $2D9 with RAPD; The Cultured cells in vitro were treated with colchicine for 6 hours, Chromosome numbers of 100 cells were examined under the microscope; The KM mice were injected intraperitoneally with 0.2ml solution of S180 - S2D9 cell, The growth of the ascitic fluid and the life span of the mice were observed; The cultured cells were fixed with 70% ethanol, DNA analysis by Flow Cytomet_ry. Results： Compared with DNA Finger map of the first generation, 50th generation, The DNA Finger map of the 100th generation had difference; the average numbers of chromosomes of 50th generation, 100th generation. 100th generation clone strain 1,100th generation clone strain2, 100th generation clone strain3, anova showed no significantly difference （P 〉 0.05） .The KM mice were injected intraperitoneally with 0.2mL solution of 5180- $2D9 cell, Ascites of three strains were bloody, The survival days changed between 13 ～ 23d, 13 ～ 20d, 10 ～ 14d, The anova showed significantly difference in 100th generation, as compared with those of the first generation and 50th generation（ P 〈 0.05）. DNA contents assayed by flow cytometry showed that DNA corresponding content of the cells is individually 0. 3575,0. 3984, 0. 3542, 0. 3919,0. 3890. Conclusion：There were not different markedly in Karyological Characteristics and DNA contents, these results of DNA Finger map and Lethality to mice hinted that the S180-S2D9 passage cells have had variance.