ANALOGS OF GUANINE NUCLEOSIDE TRIPHOSPHATES FOR SEQUENCING APPLICATIONS

We have synthesized more than 30 different deoxyribonucleosides and triphosphates with modifications either in the base or the phosphate moiety as analogs of 2′-dGTP for DNA sequencing applications. All the modified nucleoside triphosphates were tested as substrates for DNA polymerases, including Sequenase? T7 DNA polymerase or Thermo Sequenase? DNA polymerase. Two of the analogs, 7-ethyl-7-deaza-dGTP and 7-hydroxymethyl-7-deaza- dGTP meet our requirements as better sequencing reagents.     

EFFECT OF GTP AND OTHER NUCLEOSIDE TRIPHOSPHATES ON GMP INCORPORATION BY ISOLATED CORTICAL NUCLEI

—The concentration of GTP was found to be critically important in determining the characteristics of incorporation of GMP by DNA-dependent RNA polymerase from rat brain nuclei. The linearity of the incorporation rate was related to the log of the GTP concentration. Three hundred μM-GTP in the presence of the other nucleoside triphosphates (1 mM) was near to the optimal conditions in terms of maximum incorporation and linearity. The concentration of ammonium sulfate was an important factor in determining the maximum GMP and UMP incorporation. The U/G incorporation ratio was less than one at low concentrations of substrate and increased with increasing substrate or ammonium sulfate concentration. α-Amanitin strongly inhibited the reaction, indicating that RNA polymerase II is the effective enzyme.     

A STUDY OF THE NUCLEOSIDE TRI- AND DIPHOSPHATE ACTIVITIES OF RAT LIVER MICROSOMES

Rat liver microsomes catalyze the hydrolysis of the triphosphates of adenosine, guanosine, uridine, cytidine, and inosine into the corresponding diphosphates and inorganic orthophosphate. The activities are stimulated by Na₂S₂O₄, and inhibited by atebrin, chlorpromazine, sodium azide, and deaminothyroxine. Sodium deoxycholate inhibits the ATPase activity in a progressive manner; the release of orthophosphate from GTP and UTP is stimulated by low, and inhibited by high, concentrations of deoxycholate, and that from CTP and ITP is unaffected by low, and inhibited by high, concentrations of deoxycholate. Subfractionation of microsomes with deoxycholate into ribosomal, membrane, and soluble fractions reveals a concentration of the triphosphatase activity in the membrane fraction. Rat liver microsomes also catalyze the hydrolysis of the diphosphates of the above nucleosides into the corresponding monophosphates and inorganic orthophosphate. Deoxycholate strongly enhances the GDPase, UDPase, and IDPase activities while causing no activation or even inhibition of the ADPase and CDPase activities. The diphosphatase is unaffected by Na₂S₂O₄ and is inhibited by azide and deaminothyroxine but not by atebrin or chlorpromazine. Upon fractionation of the microsomes with deoxycholate, a large part of the GDPase, UDPase, and IDPase activities is recovered in the soluble fraction. Mechanical disruption of the microsomes with an Ultra Turrax Blender both activates and releases the GDPase, UDPase, and IDPase activities, and the former effect occurs more readily than the latter. The GDPase, UDPase, and IDPase activities of the rat liver cell reside almost exclusively in the microsomal fraction, as revealed by comparative assays of the mitochondrial, microsomal, and final supernatant fractions of the homogenate. The microsomes exhibit relatively low nucleoside monophosphatase and inorganic pyrophosphatase activities, and these are unaffected by deoxycholate or mechanical treatment. Different approaches toward the function of the liver microsomal nucleoside tri- and diphosphatases are reported, and the possible physiological role of the two enzymes is discussed.     

ACCURATE MASS ANALYSIS OF PHOSPHORAMIDITES BY ELECTROSPRAY MASS SPECTROMETRY

A method of accurate mass determination of phosphoramidites is described. The commonly used methanol/water/acid system was replaced by LiCl-containing acetonitrile and the concentrations of LiCl, poly(ethylene glycol), and phosphoramidite samples were optimized.     

A COMPARATIVE STUDY OF THE MORPHOLOGY AND ANATOMY OF LEUCOSTEGIA AND ARAIOSTEGIA

The anatomy and morphology of Leucostegia immersa (Wall.) Presl (type species of the genus Leucostegia) and Araiostegia pulchra (Don.) Copel. (a species which comes very near to the type species of the genus Araiostegia) are described in detail for the first time in order to determine whether the genus Araiostegia is distinct from Leucostegia. The present investigation reveals that Leucostegia differs from Araiostegia in many significant morphological and anatomical details. For example, in Leucostegia , both scales and hairs protect the rhizome, while on the rhizome of Araiostegia hairs are absent. Further, the basifixed scales of Leucostegia are sharply different from the strongly peltate scales of Araiostegia. Though the stelar cylinders in the two genera are basically similar in construction, they are markedly different in their modes of production of root-traces and leaf-traces. No tanniniferous cells which are found in Leucostegia , occur in the fundamental tissues of Araiostegia. Young fronds of Leucostegia produce delicate hairs, but in Araiostegia the fronds are glabrous throughout. The rachises and costae in Leucostegia are grooved on the upper surface and never show any raised median bands in sharp contrast to the rachises of Araiostegia which are at least in the upper part narrowly winged on either side of the raised upper surface. All these differences certainly show that these genera are well defined. The relationship between Araiostegia and other davallioid ferns is also discussed.     

MONO‐ AND DIGALACTOSYLDIACYLGLYCEROL COMPOSITION OF RAPHIDOPHYTES (RAPHIDOPHYCEAE): A MODERN INTERPRETATION USING POSITIVE‐ION ELECTROSPRAY/MASS SPECTROMETRY/MASS SPECTROMETRY1

Raphidophyte algae (Raphidophyceae) can be divided according to pigment composition and plastid ancestry into two categories, brown‐ and green‐pigmented taxa. We sought to examine if there are any biochemical differences in plastid lipid composition between the two groups. To this end, the composition and positional distribution of fatty acids of the chloroplast lipids, mono‐ and digalactosyldiacylglycerol (MGDG and DGDG, respectively), were examined using positive‐ion electrospray/mass spectrometry (ESI/MS) and electrospray/mass spectrometry/mass spectrometry (ESI/MS/MS). Brown‐pigmented strains from the genera Chattonella, Fibrocapsa, and Heterosigma primarily consisted of 20:5/18:4 (sn‐1/sn‐2) MGDG and 20:5/18:4 DGDG, while isolates of the green‐pigmented raphidophyte Gonyostomum semen (Ehrenb.) Diesing contained these as well as 18:3/18:4 MGDG and DGDG, thus underscoring its green algal plastid lineage. Although previously unseen without the regiochemical information provided by ESI/MS/MS, Chattonella subsalsa Biecheler possessed 20:5/18:3 DGDG as a major form, a potential biosynthetic intermediate in the production of 20:5/18:4 DGDG. These results provide a modern interpretation of the fatty acid regiochemistry of MGDG and DGDG.     

环烯醚萜甙的快速原子轰击质谱和场解吸质谱研究

环烯醚萜甙是植物界分布较广泛的一类配糖体成分,极性较大,用通常的电子轰击质谱测试不能得到分子离子信息,且离子碎片零碎,难以进行结构解析。本文对２５个不同类型的环烯醚萜甙进行正,负离子快速原子轰击质谱和场解吸质谱分析,并讨论这两种质谱新技术在环烯醚萜甙类化合物结构解析中的应用。     

LASER MICROPROBE MASS SPECTROMETRY: PRINCIPLE AND APPLICATIONS IN BIOLOGY AND MEDICINE

Laser microprobe mass spectrometry (LMMS) is an interesting technique for micro- and surface analysis. It employs local ionization by a focused laser under high power density conditions and subsequent mass analysis of the generated ions. This paper surveys the main LMMS instruments and their operational principles. Sample preparation is discussed in the context of biological materials. The problem of quantification is addressed. Selected examples show the way that precise information on the molecular composition can be deduced from the detected signals. Both inorganic and organic substances can be identified, even without reference spectra, from in-situ analysis with a lateral resolution in the order of 1 to 5μm.     

A COMPARATIVE STUDY OF THE OLFACTORY SENSITIVITY OF HUMANS AND RATS

Olfactory sensitivity of humans and rats to n-propanol, n-heptanol,benzaldehyde, isobutyln-butyrate, cyclohexanone and 1,4-dioxane(rats only), was determined. Similar results were obtained forboth species, and these compare favourably with those of otherworkers. Since the sensitivities of humans and rats varied fordifferent odorants in a similar manner, it is postulated thatthe mechanisms for detecting odors may be common to many vertebrates.It is proposed, in agreement with Adrian (1956), and Moultonet al. (1960), that the relative area of the olfactory epitheliumdoes not necessarily determine the sensitivity of an animal. ^*This paper reports research undertaken at the School of BiologicalSciences, Macquarie University, North Ryde, N.S.W., 2113, Australia,and the CSIRO Division of Food Research.     

A COMPARATIVE STUDY OF PINEAL ACETYL-CoA HYDROLASE AND N-ACETYLTRANSFERASE ACTIVITIES

pineal acetyl-CoA hydrolase is measurable at 4 days before birth. It increases rapidly to a maximum of 0.37 nmol/min/0.1 mg protein during the first week after birth, thereafter gradually decreasing and stabilizing at adult levels (0.27 nmol/min/0.1 mg protein) 3-4 weeks after birth. Unlike A/-acetyltransferase, the activity of acetyl-CoA hydrolase does not increase following treatment with isoproterenol, does not exhibit a circadian rhythm and is not inactivated on exposure of the animals to light at night. In addition, denervation of the pineal gland does not alter acetyl-CoA hydrolase activity.     

红豆草和苜蓿的光合效率比较研究

红豆草,又名驴食草,是甘肃省近年广泛种植的豆科牧草。为了深入认识它的生长发育规律,揭示其光合生产的限制因素,我们于1986—1988连续三年在甘肃省陇中南部黄土丘陵区通渭县申家山试验场开展了红豆草光能利用率的研究。