LOCALIZATION OF STARCH BIOSYNTHETIC AND DEGRADATIVE ENZYMES IN MAIZE LEAVES

The cellular distribution of the starch biosynthetic and degradative enzymes in protoplasts prepared from maize leaf mesophyll and bundle sheath cells was investigated. In conformity with the cellular distribution of starch, starch biosynthetic enzymes (soluble starch synthase, ADPglucose pyrophosphorylase, branching enzyme and starch Phosphorylase) were exclusively localized in the bundle sheath cells. In contrast, starch degradative enzymes (α-amylase, β-amylase and debranching enzyme) were present in both types of leaf cells. Isolated chloroplasts from bundle sheath cells were shown to contain 100% of the starch biosynthetic enzymes. However, approximately 60% of the activity of degradative enzymes and 67% of the activity of starch Phosphorylase was localized in bundle sheath chloroplasts.     

甘蓝型油菜子叶和下胚轴再生植株无性系建立

以甘蓝型油菜(Brassica napus L.)豫油2号和6257的子叶和下胚轴为材料,在不同激素配比的MS培养基上诱导出了愈伤组织。将经过继代的部分愈伤转入分化培养基,分化结果表明:除基因型、外植体和分化培养基的激素配比对分化率有影响外,诱导愈伤培养基的激素配比对分化率也至关重要。豫油2号的子叶和下胚轴在最适诱导培养基(ZT 1+NAA0.5+2,4-D 0.2 mg/L)和最适分化培养基(ZT4+IAA 0.2 mg/L)组合中的愈伤分化率分别为12.5%和75%;6257的子叶和下胚轴在其最适诱导培养基(KT 2+NAA1+2,4-D 0.2 mg/L)和最适分化培养基(6-BA 4+IAA 0.02 mg/L)组合中的愈伤分化率分别为50%和37.5%。将其最适诱导培养基中的愈伤组织继代达8个月以上,建立了不同继代愈伤的再生植株无性系。     

STARCH GEL ELECTROPHORESIS OF ENZYMES FROM NINE SPECIES OF POLYPORUS

Taxonomic relationships among 22 isolates of nine species of the wood-rotting fungus, Polyporus, were examined by starch gel electrophoresis, coupled with histochemical staining procedures. Crude extracts of 21-day cultures grown on malt extract agar were subjected to electrophoresis for 3 hr at a constant current of 25 ma. Esterase, peroxidase, catechol oxidase, galactose dehydrogenase, acid phosphatase, and alkaline phosphatase banding patterns were analyzed. With each enzyme, the bands for each isolate were compared with those for other isolates of the same species and for isolates of other species. A closer relationship was found in the banding patterns for different isolates of a single species than among isolates of different species. This technique may prove valuable for the evaluation of taxonomic differences among these wood-rotting fungi.     

发根土壤杆菌体外转化甘草子叶及下胚轴

利用发根土壤杆菌农杆碱型15834、A_4菌株和甘露碱型8196、K_(599)菌株体外转化甘草子叶及下胚轴。结果表明:除K_(599)菌株外,其余三种菌株都能诱导产生甘草发状根无性系。其中15834菌株的诱导率最高,下胚轴的诱导率高于子叶。组织学观察表明,在感染后4天左右,下胚轴维管束鞘外细胞、特别是形成层部分细胞大量启动形成分生细胞及分生细胞团,并进一步分裂形成根原基。6—8天后根原基具单向极性生长形成完整的发状根结构。发状根能在无外源激素的培养基上迅速生长。高压纸电泳检测到发状根中存在农杆碱及甘露碱,说明Ri质粒T-DNA编码的这两种冠瘿碱合成酶基因在甘草细胞中得到了表达。     

SOME ENZYMES RELATING TO THE METABOLISM OF STARCH IN A SEA-LETTUCE

The occurrence of a phosphorylase in the extracts of a greenseaweed, Ulva pertusa was demonstrated. In addition, a sugarphosphatase different from non-specific acid phosphatase andan amylase, which are involved in the starch degradation, werealso detected in this sea-lettuce. ¹Contributions from the Shimoda Marine Biological Station, No.147.     

LOCALIZATION OF THE ENZYMES OF KETOGENESIS IN RAT LIVER MITOCHONDRIA

The localization of the enzymes of ketogenesis in isolated rat liver mitochondria has been investigated. Mitochondrial subfractions were isolated after disruption of this subcellular organelle by (a) hypotonic lysis in water, which permitted the ultracentrifugal separation of the soluble and membranous compartments of the mitochondrion, or by (b) a procedure involving swelling, contraction, and ultrasonic treatment, which permitted the isolation from discontinuous sucrose gradients of subfractions rich in intermembrane space protein, outer membrane, and inner membrane-matrix particles. Two membrane subfractions were invariably present as distinct bands at the lower interface of the discontinuous gradient. The upper of these two bands was found to be a highly purified preparation of outer mitochondrial membrane. Subfractions rich in matrix and in inner membrane were isolated from inner membrane-matrix particles after hypotonic treatment. The content of the various mitochondrial compartments in all subfractions was assessed from their enzymic and electron microscopic characteristics. The ketogenic activity of each subfraction was determined by measuring its capacity to form ketone bodies from acetyl CoA. The activity of this process was markedly enhanced by dithiothreitol. These measurements of ketone body formation, together with assays of individual enzymes of the ketogenic pathway, show that thiolase, HMGCoA synthase, and HMGCoA cleavage enzyme are localized in the matrix of the inner membrane-matrix particles. The rates of ketone body formation indicate that the HMGCoA synthase is the rate-limiting enzyme of the pathway in subfractions of high matrix content. Studies with sodium chloride indicate that a large portion of the HMGCoA synthase, which remains present in membrane subfractions derived from water-treated mitochondria, is bound by ionic interaction to component(s) of the membrane.     

蒜鳞片薄壁细胞衰退过程中酶的细胞化学定位及DNA生化分析

蒜在储藏过程中,鳞茎薄壁细胞衰退,其营养物质供给幼芽萌发生长。采用细胞化学方法,对蒜休眠进程中的鳞茎薄壁细胞进行了ATPase以及APase的细胞化学定位,结果显示在薄壁细胞的质膜、细胞壁和胞间连丝上的酶活性随着蒜自休眠至萌发的不同发育进程而呈现增强的趋势,且在萌芽期酶活性表现最为强烈,表明细胞内物质的降解、转化与输出的加强有助于细胞内含物向新生芽的彻底转移。配合采用琼脂糖凝胶电泳对衰退薄壁细胞的DNA进行了分析,实验结果表现出典型的DNA Ladder,为蒜鳞茎薄壁细胞的衰退属于受基因控制的程序性死亡范畴补充了生化证据。     

蒜鳞片薄壁细胞衰退过程中酶的细胞化学定位及DNA生化分析

蒜在储藏过程中。鳞茎薄壁细胞衰退。其营养物质供给幼芽萌发生长。采用细胞化学方法,对蒜休眠进程中的鳞茎薄壁细胞进行了ATPase以及APase的细胞化学定位,结果显示在薄壁细胞的质膜、细胞壁和胞间连丝上的酶活性随着蒜自休眠至萌发的不同发育进程而呈现增强的趋势,且在萌芽期酶活性表现最为强烈。表明细胞内物质的降解、转化与输出的加强有助于细胞内含物向新生芽的彻底转移。配合采用琼脂糖凝胶电泳对衰退薄壁细胞的DNA进行了分析,实验结果表现出典型的DNA Ladder．为蒜鳞茎薄壁细胞的衰退属于受基因控制的程序性死亡范畴补充了生化证据。     

等位酶淀粉凝胶电泳技术中的几个应引起重视的问题

本文总结了我们在用淀粉凝胶电泳技术进行等位酶分析的实验过程中的一些经验和教训,主要结论是：１）尽管材料不同,对于不同的酶系统,用特定的缓冲液系统常能获得比较理想的效果,如ＤＩＡ,ＧＰＩ,ＨＥＸ,ＳＯＤ和ＴＰＩ用Ｓ６（Ｓｏｌｔｉｓ等,１９８３）;ＡＭＰ用Ｓ８;ＭＤＨ用Ｓ９和ＳＫＤ用Ｗ２（Ｗｅｎｄｅｌ和Ｗｅｅｄｅｎ,１９８９）等。２）用磷酸缓冲液配制ＡＭＰ的染色配方较其它配方效果好。３）当胶染时,使用琼脂替代琼脂糖,染色效果不变而成本更低,谱带扩散更慢。４）不同的提取液可能适用于不同的酶系统,如磷酸ＰＶＰ提取液（Ｓｏｌｔｉｓ等,１９８３）不适用于石荠苎的ＧＰＩ的提取。５）正确选择凝胶和电极缓冲液,Ｓ１虽然适用面广,但效果不一定好,对于位点多和等位基因丰富的材料尤其应警惕,因Ｓ１可能掩盖样品之间的差异。     

EFFECTS OF S-TRIAZINES ON PROTEIN AND FINE STRUCTURE OF COTYLEDONS OF BUSH BEANS

A significant increase in protein content of bean cotyledons resulted by applications of 0.5 ppm of atrazine (2-chloro-4-ethylamino-6-isopropylamino-s-triazine), simazine [2-chloro-4,6-bis(ethylamino)-s-triazine], terbutryn (2-methylmercapto-4-ethylamino-6-isobutylamino-.s-triazine), or GS-14254 (2-methoxy-4-isopropylamino-6-butylamino-s-triazine) to the foliage of 5-6 week old bean plants grown in a controlled environment or field conditions. Aen electron microscopic study indicated that in the cotyledonary cells s-triazines inducd a 2-fold increase in the number of cisternae of rough endoplasmic reticulum. These treatments also increased the number of vesicles, which apparently contain protein, and the amount of cytoplasmic ribosomes.