CELLULAR FRACTIONATION OF CHLAMYDOMONAS-REINHARDII WITH EMPHASIS ON THE ISOLATION OF THE CHLOROPLAST
CELLULAR FRACTIONATION OF CHLAMYDOMONAS-REINHARDII WITH EMPHASIS ON THE ISOLATION OF THE CHLOROPLAST
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DOI：
10.1104/pp.72.2.481
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发表时间：
1983-01-01
期刊：
影响因子：
7.4
通讯作者：
PLATTALOIA, KA
中科院分区：
文献类型：
--
作者：
KLEIN, U;CHEN, CG;PLATTALOIA, KA
本文描述了一种从藻的同步细胞中分离出完整叶绿体的方法。细胞分离过程主要包括4个步骤：用自溶酶生产原生质体，用洋地黄苷裂解原生质体，破碎原生质体聚集，用差速离心分离细胞器。用Percoll缓冲离心代替差速离心，得到完整的叶绿体。从100毫升海藻培养液开始，培养液中含有3000。G叶绿素，叶绿体完整，100-200亩。叶绿素的G可以被分离出来。包膜完整性约为90%（铁氰化物测定）。叶绿体的电镜和标记酶活性检测表明线粒体和细胞质受到污染。
A method for cellular fractionation of C. reinhardii, SAG 11-32/b, and isolation of intact chloroplasts from synchronized cells of the alga is described. The procedure for cell fractionation comprises essentially 4 steps: protoplast production with autolysine, lysis of the protoplasts with digitonin, aggregation of broken protoplasts, and separation of organelles by differential centrifugations. Replacing the differential centrifugations by Percoll cushion centrifugations yields intact chloroplasts. Starting with 100 ml of an algal culture containing 3000 .mu.g chlorophyll, intact chloroplasts with 100-200 .mu.g of chlorophyll can be isolated. Envelope integrity is about 90% (ferricyanide assay). Examination of the chloroplasts by EM and marker enzyme activities indicated some mitochondrial and cytoplasmic contamination.