Studies on the Analysis of Fluorescence Decay Data by the Method of Moments

The method of moments, as presented by Isenberg and Dyson (1969; Biophys. J. 9:1337) has been shown to be a reliable way of obtaining the amplitudes and time constants of several simultaneously emitting species, even in the presence of an overlapping excitation. Recent improvements in the method include (a) a component incrementation test for determining the number of relaxations, (b) a procedure, which we call exponential depression, for dramatically improving convergence, and (c) a new algorithm for implementing the method of moments on a digital computer with a high degree of flexibility and efficiency. These improvements, as well as new general theory, are described and tested using both synthetic and real experimental data. Component incrementation consists of examining models with increasing numbers of exponential terms. Given adequate precision, we find that an analysis for N + 1 components, of data that are actually represented by N components, provides the correct amplitudes and time constants plus an N + 1 term with an insignificant amplitude. Exponential depression is a transformation in which the original excitation and fluorescence, E(t) and F(t), are multiplied by exp (-λt), where λ is an arbitrary parameter. While the convolution is invariant to this transformation, the proper choice of λ greatly reduces the number of iterations necessary to obtain the amplitudes and time constants and may even improve their accuracy. In addition, an appendix by John P. Mullooly presents a statistical analysis of the effect of counting error on the method of moments estimates of fluorescence decay parameters, applicable when data are obtained by the monophoton technique. Formulas are derived that give the approximate precision of the decay parameters for the general case of N exponential components, with calculational details for one and two component systems.     

The circular dichroism of poly d(AAT): d(ATT) and poly r(AAU):r(AUU). A test of the first-neighbor hypothesis

We present absorption and circular dichroism (CD) spectra for the synthetic polymers poly d(AAT):d(AAT) and poly r(AAU):r(AAU), in both native and heat-denatured forms. As a means of evaluating the first-neighbor hypothesis, the CD spectra are compared with approximations derived from spectra of other synthetic polymers containing the same first-neighbor sequences. This is the first instance where such a comparison has been possible using spectra of double-stranded RNA sequences, and the agreement between the measured and approximated spectra for poly r(AAU):r(AUU) is surprisingly good. We have also subjected the CD spectrum of poly d(AAT):d(AAT) to a previously published analytical procedure for obtaining estimates of first-neighbor frequencies. In this first independent test of the procedure, we find that the analysis does infer the existence of a majority (86%) of AA, TT, AT, and TA first neighbors but does not precisely indicate their relative proportions.     

Studies of nonmigrating long-lived lymphocytes in rats. V. Presence and reactions to antigens after neonatal thymectomy

Neonatally thymectomized and sham-operated rats were given courses of hind foot pad injections of tritiated thymidine after immune stimuli, and the popliteal lymph nodes were examined for the presence of persistently labeled lymphocytes 32 days later. A larger proportion of labeled small lymphocytes was found in the nodes of thymectomized rats than in the nodes of sham-operated rats, indicating that a significant portion of the long-lived nonmigrating cells are B cells. In sham-operated rats given a course of tritiated thymidine after an immune stimulus with a Salmonella flagellin, labeled blasts appeared 3 days after secondary challenges with both the same or a serologically distinct flagellin. In neonatally thymectomized rats, a blastogenic response occurred after challenge with the same flagellin but not after challenge with the non-specific flagellin. Thus, in normal rats there are also long-lived nonmigrating T cells in primed lymph nodes, and these are the cells responsible for the nonspecific blasto-genesis seen in earlier experiments.     

Comportement des chromosomes surnuméraires et relation avec le taux de mortalité dans une population africaine de Locusta migratoria migratorioides

Transmission of supernumerary chromosomes is studied in adults and embryos of an African population of Locusta migratoria migratorioides; 34% of the animals show one or two B chromosomes (24% one, 10% two). This percentage is the same in both sexes. During mitosis, B chromosomes are very stable. At meiosis, in some cases they show pairing, in other cases they enter as univalents. The eventuality of synapsis or association is discussed. These two kinds of behaviour could not be explained by the presence of two different B chromosomes as is shown from the study of parthenogenetic progeny. Repartition of B chromosomes in the progeny is different depending on whether the male or the female parent supplies them; so the behaviour of these supernumerary should be conditioned, not by their own structure, but by a connection with the cell. — One B chromosome apparently is neutral. When there are two B chromosomes, in case of synapsis, the number of animals with two B in the progeny is slightly lower than previous; in case of asynapsis, there is a higher lethality of individuals and oocytes with two B chromosomes.     

Determination of Melting Sequences in DNA and DNA-Protein Complexes by Difference Spectra

A graphical formula is presented for determining the base ratio of melted DNA. By use of this formula, the composition of sequences which melt in different portions of the melting curves of Clostridium DNA, Escherichia coli DNA, and mouse DNA were determined. As the DNA melts, the per cent of adenine and thymine (AT) in the melted sequences decreases linearly with temperature. The average composition of sequences which melt in a given part of the melting curve is proportional to the base ratio of the DNA. The concentration and average composition of sequences were determined for three parts of the melting curves of the DNA samples, and a frequency distribution curve was constructed. The curve is symmetrical and has a maximum at about 56% AT. The distribution of GC-rich sequences on the E. coli chromosome was estimated by shearing, partially melting, and fractionating the DNA on hydroxylapatite. GC-rich sequences appear to occur every thousand base pairs, and have a maximum length of about 180 base pairs. The graphical formula was applied to the determination of the composition of sequences which melt in different parts of the melting curve of chromatin. Throughout the melting curve, the composition of the melting sequences is about 60% AT, which appears to suggest that relatively long sequences are melting simultaneously. Their melting temperature may be a function of the composition of the protein on different parts of the DNA. The problem of light scattering in DNA-protein and DNA was also investigated. A formula is presented which corrects for light scattering by relating the intensity of the scattered light to the rate of change of absorbance of DNA with wavelength.     

A new Costa Rican species of centropogon (Campanulaceae-Lobelioideae)

A newCentropogon (sect.Wimmeriopsis subsect.Colombiani) from the Cordillera de Talamanca, Costa Rica, is described and compared toC. gutierrezii (Planch. & Oerst.) F. E. Wimmer, a species restricted to the Cordillera Central with which it previously has been confused.     

SEEDLING MORPHOLOGY IN MARAH (CUCURBITACEAE) RELATED TO THE CALIFORNIAN MEDITERRANEAN CLIMATE

Studies in reproductive ecology were made in indigenous, western American plants in the genus Marah (Cucurbitaceae), with particular attention given plants of M. oreganus occurring in the Berkeley Hills near San Francisco Bay in California. These tuberous perennials produce capsular fruits on their annual aboveground shoots; the fruits dehisce in early summer, each one exposing about three large seeds with an average seed weight of 1.05 g. The embryo of a M. oreganus seed has two thick and fleshy cotyledons packed with protein granules. The embryonic axis, with shoot and root apices, is ca. 0.5-1.0 mm long, roughly ½0 or less the length of the seed. In the Berkeley Hills dispersal of the seeds is accomplished by nocturnal rodents, after which germination begins with the fall rains and cooler temperatures of November and December. Instead of a radicle emerging first from the seed at germination, the minute radicle and epicotyl are pushed or carried far out of the seed, down into the soil, by the elongating bases of the cotyledons. These cotyledon bases, or petioles, are fused, and as they elongate they form a hollow tube that bears the embryonic axis at its extreme tip. The cotyledonary petiole tube ceases elongation by January, when it may be 5-25 or more cm long in a seedling of M. oreganus. Then, from its tip, the radicle grows downward and the epicotyl upward—up the hollow petiole tube. The green shoot (epicotyl) reaches the soil surface by early March in this area, completes the first season's growth, and dries up by late May, when the arid summer season is beginning. But even before the epicotyl grows out of the petiole tube and above ground, the seedling's hypocotyl begins to enlarge, forming a tuber. The fleshy cotyledon blades remain in the seed coat below ground, and some food from the blades is transferred to the tuber that produces shoots in the following growing seasons. This pattern of germination and seedling establishment is now known for species of Marah and for a very few other dicotyledonous plants, all of them growing mainly in areas of hot and dry habitat that are generally referred to as having Mediterranean climate. This elongation of the fused hypogeal cotyledons is considered a complex adaptation in dicotyledons that helps ensure fast and successful seedling establishment in seasonally arid areas such as “Mediterranean” California.