杨树新品种叶肉原生质体培养和植株再生

从１ 个月龄的ＮＬ－８０１０６ 杨（Ｐｏｐｕｌｕｓｄｅｌｔｏｉｄｅｓ×Ｐ． ｓｉｍ ｏｎｉｉ）无菌苗叶片分离得到大量原生质体,纯化后其原生质体产量为４×１０７／ｇ ｆｒ．ｗ ｔ． 纯化的原生质体在含２,４－Ｄ ２ ｍ ｇ／Ｌ、ＮＡＡ ０．５ ｍ ｇ／Ｌ和ＫＴ ０．５ ｍ ｇ／Ｌ的ＫＭ８ｐ 和ＭＳ培养基中进行高密度液体浅层培养,渗透势为０．４０ ｍ ｏｌ／Ｌ的ＫＭ８ｐ 培养基中原生质体分裂频率最高． 培养第５ 天观察到第一次细胞分裂,培养１０ ｄ 的分裂频率为４．５％ ,１２ 周内可形成大量的细胞团和小愈伤组织． ＮＬ－８０１０６杨叶肉原生质体在富含有机氮并以葡萄糖为碳源的培养基中具有较高的分裂频率和植板率．小愈伤组织在ｇｅｌｒｉｔｅ 固化的ＮＬＺ１ 培养基上增殖生长,３ 周后形成４—６ ｍ ｍ 结构紧密的鲜红色愈伤组织,转至ＮＬＦ分化培养基,分化成苗率为１００％ ． 待芽伸长到３ ｃｍ 时,从基部切下转至１／２ ＭＳ培养基上诱导生根,形成完整植株     

Crystallization,molecular replacement solution,and refinement of tetrameric β-amylase from sweet potato

Sweet potato β-amylase is a tetramer of identical subunits, which are arranged to exhibit 222 molecular symmetry. Its subunit consists of 498 amino acid residues (Mr 55,880). It has been crystallized at room temperature using polyethylene glycol 1500 as precipitant. The crystals, growing to dimensions of 0.4 mm × 0.4 mm × 1.0 mm within 2 weeks, belong to the tetragonal space group P4₂2₁2 with unit cell dimensions of a = b = 129.63 Å and c = 68.42 Å. The asymmetric unit contains 1 subunit of β-amylase, with a crystal volume per protein mass (V_M) of 2.57 ų/Da and a solvent content of 52% by volume. The three-dimensional structure of the tetrameric β-amylase from sweet potato has been determined by molecular replacement methods using the monomeric structure of soybean enzyme as the starting model. The refined subunit model contains 3,863 nonhydrogen protein atoms (488 amino acid residues) and 319 water oxygen atoms. The current R-value is 20.3% for data in the resolution range of 8–2.3 Å (with 2 σ cut-off) with good stereochemistry. The subunit structure of sweet potato β-amylase (crystallized in the absence of α-cyclodextrin) is very similar to that of soybean β-amylase (complexed with α-cyclodextrin). The root-mean-square (RMS) difference for 487 equivalent Cα atoms of the two β-amylases is 0.96 Å. Each subunit of sweet potato β-amylase is composed of a large (α/β)₈ core domain, a small one made up of three long loops [L3 (residues 91–150), LA (residues 183–258), and L5 (residues 300–327)], and a long C-terminal loop formed by residues 445–493. Conserved Glu 187, believed to play an important role in catalysis, is located at the cleft between the (α/β)₈ barrel core and a small domain made up of three long loops (L3, L4, and L5). Conserved Cys 96, important in the inactivation of enzyme activity by sulfhydryl reagents, is located at the entrance of the (α/β)₈ barrel. © 1995 Wiley-Liss, Inc.     

Frequency analysis of time-varying elastance model of the left ventricle

Zadeh's transfer function method for linear time-variable systems is used to apply frequency-domain analysis to a periodically time-varying elastance model of the left ventricle. Left ventricular pressure computed from the system function of the time-varying elastance and the phasors of aortic flow shows a typical waveform of the measured ventricular pressure.     

Bacteriophage MS2 RNA: A correlation between the stability of the codon: Anticodon interaction and the choice of code words

Summary The non-random distribution of degenerate code words in Bacteriophage MS2 RNA can be explained partially by considerations of the stability of the codon-anticodon complex in prokaryotic systems. Supporting this hypothesis we note that wobble codons are positively selected in codons having G and/or C in the first two positions. In contrast, wobble codons are statitically less likely in codons composed of A and U in the first two positions. Analyses of nucleotides adjacent to 5 and 3 ends of codons indicate a nonrandom distribution as well. It is thus likely that some elements of RNA evolution are independent of the structural needs of the RNA itself and of the translated protein product.This work was supported by grants from the Belgian Government Actions concertées - Gekon-certéerde Acties, N.F.W.O. and F.K.F.O. as well as from le Ministère de l'éducation du Québec. A preliminary report of this work was given at the EMBO ribosome workshop, Brussels 1976     

Autoregulation of the stability operon of IncFII plasmid NR1.

The stb locus of IncFII plasmid NR1, which mediates stable inheritance of the plasmid, is composed of an essential cis-acting DNA site located upstream from two tandem genes that encode essential stability proteins. The two tandem genes, stbA and stbB, are transcribed as an operon from promoter PAB. Using PAB-lacZ gene fusions, it was found that the stb operon is autoregulated. A low-copy-number stb+ plasmid introduced into the same cell with the PAB-lacZ fusion plasmid repressed beta-galactosidase activity about 5-fold, whereas a high-copy-number stb+ plasmid repressed beta-galactosidase about 15-fold. The details of autoregulation were analyzed by varying the concentrations of StbA and StbB to examine their effects on expression from the PAB-lacZ fusion plasmid. StbB protein by itself had autorepressor activity. Although StbA protein by itself had no detectable repressor activity, plasmids that encoded both stbA and stbB repressed more effectively than did those that encoded stbB alone. Plasmids with a mutation in stbA had reduced repressor activity. One mutation in stbB that inactivated the stability function also reduced, but did not eliminate, repressor activity. Repressor activity of the mutant StbB protein was effectively enhanced by stbA. These results indicate that StbB serves two functions, one for stable inheritance and one for autoregulation of the stb operon, both of which may be influenced by StbA protein.     

Purification and some properties of an extracellular l-amino acid oxidase from Cellulomonas cellulans AM8 isolated from soil

Summary The soil isolate Cellulomonas cellulans AM8 produces an extracellular l-amino acid oxidase (L-AAO) with broad substrate specificity. The strain produced up to 0.35 unit (U)/ml of the extracellular L-AAO in a simple medium containing glycerol and yeast extract. The enzyme was easily purified up to 30 U/mg protein using Phenyl-Sepharose fast flow. The purified enzyme migrated as single band on sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) with a molecular mass of 55 kDa. On native PAGE the molecular mass was approx. 300 000 kDa, which may be due to aggregation. With the exception of glycine, proline, and threonine, all the amino acids normally constituting proteins were oxidized. The V _max values from 0.7 to 35.2 U/mg for aspartic acid and lysine, respectively, and the K _m values from 0.007 to 7.1 mm for cysteine and valine, respectively, were obtained at 25° C and pH 7.0 in oxygen-saturated solutions. The L-AAO had a pH optimum of 6.5–7.5. It was stable for several months at — 30° C and for some days at 35° C. Ferricyanide served as an electron acceptor with a V _max of 50 U/mg and K _m for 0.3 mm with phenylalanine as the substrate. Correspondence to: R. D. Schmid     

Determination of an antisecretory trimethyl prostaglandin E2 analog in human plasma by combined capillary column gas chromatography—negative chemical ionization mass spectrometry

A method is described for measuring a trimethyl prostaglandin E₂ analog, TM-PGE₂, in human plasma. Trideuterated and monofluorinated analogs of TM-PGE₂ are added to plasma as internal standard and carrier, respectively. The plasma is adjusted to pH 3.0 and is extracted with a mixture of benzene—dichloromethane (9:1). The residue, following removal of the extracting solvent, is reacted consecutively with pentafluorobenzyl bromide and bistrimethylsilyltrifluoroacetamide. The excess derivatizing reagents are removed by evaporation, and an aliquot of the reconstituted residue is analyzed by capillary column gas chromatography using methane as the carrier gas. A quadrupole mass spectrometer is set to monitor in the gas chromatographic effluent the (M − C₇H₂F₅)⁻ fragmention of TM-PGE₂ (m/e 449) and trideuterated TM-PGE₂ (m/e 452) generated by methane negative chemical ionization. Quantitation of unknowns is based on a comparison of the m/e 449 to m/e 452 ion ratio in each unknown to that obtained from the analysis of control plasma spiked with known amounts of TM-PGE₂ and fixed amounts of internal standard and carrier. The sensitivity limit of the assay is approximately 100 pg ml⁻¹, which is equivalent to 1 pg injected. The assay was used to measure the concentration of TM-PGE₂ in the plasma of two subjects following a single 10 μg kg⁻¹ oral dose of the drug.     

Inhibition of Ouabain-binding to (Na+ + K+)ATPase by antibody against the catalytic subunit but not by antibody against the glycoprotein subunit

Antibodies against purified ($\text{Na}{}^{\mathrm{+}}\text{+}\text{K}{}^{\mathrm{+}}\text{)}\text{ATPase}$ from the rectal gland of Squalus acanthias, as well as against its catalytic subunit, inhibited ouabain binding by as much as 50%. However, antibodies against the glycoprotein subunit did not inhibit ouabain binding. These data suggest that binding of antibody against the catalytic subunit to the enzyme either covers the ouabain binding site or destroys its conformation, while binding of antibody against the glycoprotein has no such effect.