Observations presented in this paper point to the presence of dual transport mechanisms for the base adenine in Neurospora crassa. Competition for transport, as well as growth inhibition studies using an ad-i auxotroph, show that the purine bases adenine, guanine, and hypoxanthine share at least one transport mechanism which is insensitive to adenosine, cytosine, and a variety of other purine base analogues. On the other hand, uptake of adenine by an ad-8 mutant strain unable to transport [8-14C]hypoxanthine at any concentration was not inhibited by guanine or hypoxanthine. This observation demonstrates the existence of an adenine-specific transport system which was also found to be insensitive to inhibition by other purine base analogues, adenosine or cytosine. Recombination analysis of ad-8 by wild-type crosses showed that the inability to transport [8-14C ]hypoxanthine was a consequence of the ad-8 lesion or a closely linked mutation. Saturation plots of each system gave intermediary plateaus and nonlinear reciprocal plots which, based on comparison with pure enzyme kinetic analysis, suggest that either each system consists of two or more uptake systems, at least one of which exhibits cooperativity, or that each system is a single uptake mechanism which possesses more than two binding sites where the relative affinity for the purine base first decreases and then increases as the sites are filled.

Significantly more 5-methylcytosine residues were found in the DNA from the dormant sclerotia of Phymatotrichum omnivorum than in the DNA from the metabolically active mycelia of the fungus, as shown by high-pressure liquid chromatography of acid-hydrolyzed DNA digests and by restriction of the DNA with the isoschizomers MspI and HpaII. N6-Methyladenine was not detected in GATC sequences in the DNA isolated from either stage.The occurrence of significant amounts of 5-methylcytosine (5-mG) appears to be rare in the DNA of lower eucaryotes.Antequera et al. (1) analyzed DNA isolated from mycelia of 20 species of fungi representing 15 taxonomic families and found only 2 species in which there was detectable methylation of cytosine residues, as demonstrated by comparison of MspI and HpaII restriction patterns. The same authors reported that very little 5-mC was found in mycelial DNA from Aspergillus nidulans (24). Proffitt et al. reported that less than 0.01% of the cytosine residues were methylated in the DNA of Saccharomyces cerevisiae (19). A high level of 5-mC (21%) was found in Chlorella nuclear DNA and in several Chlorella viruses, but was not found in other green algae (26). Bull and Wootton demonstrated that amplified transforming DNA from Neurospora crassa mycelia was heavily methylated, whereas the nuclear DNA from wildtype mycelia had very low levels of 5-mC based on analysis of MspI and HpaI digests (5).There is considerable evidence in higher eucaryotic organisms, especially vertebrates, that gene inactivation may be caused by methylation of specific cytosine residues in the DNA. In humans, the developmental switch from fetal to adult forms of hemoglobin (6,13,15) and the inactivation of genes on one X chromosome in females (16,30,31) are accompanied by the methylation of cytosine residues in 5'-flanking regions of the DNA. However, the lack of significant methylation of DNA from lower eucaryotes such as fungi and from some insects such as Drosophila melanogaster (25) indicates that DNA methylation is not a universal mechanism for inactivating genes during development.The base N6-methyladenine (6-mA) has been detected in the DNA of a number of lower eucaryotes including the unicellular green alga Chlamydomonas reinhardi (12), several dinoflagellates (20), ciliated protozoa (3, 7, 11), and some insects (8). Experiments with Tetrahymena thermophila (3) suggest that 6-mA may serve to protect specific DNA sequences from degradation during developmental changes in the life cycle. 6-mA has not been found in fungi and appears to be absent from the DNA of all higher eucaryotes (8).This work compared the methylation of DNA from differentiated resting structures (sclerotia) and actively growing, proliferative mycelia of the fungus Phymatotrichum omnivorum by using restriction by isoschizomers differing in * Corresponding author. sensitivity to base methylation and by using quantitation by high-pressure liquid chromatography of the 5-mC content in acid-hydrolyzed DNA. MATERIALS AND METHODSStrains and culture condit...

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Transgenic petunia (Petunia hybrida Vilm.) plants were obtained from Agrobacterium-mediated shoot apex transformation. Studies at the phenotypic as well as molecular level established both the presence of the NPT II (neomycin phosphotransferase II) and GUS (β-glucuronidase) genes and their level of activity. Twenty-nine primary transformed plants showed varying patterns of phenotype expression of both genes. NPT II and GUS expression in 7 primary plants over a 4-month interval showed varying levels of gene expression within and among individual plants. All primary transgenic plants were self-pollinated and backcrossed to establish the inheritance patterns of both genes. Mendelian and non-Mendelian inheritance patterns for both genes were observed. Analysis of the progeny showed poor transmission of the foreign genes through the pollen especially when two or more bands were present in the Southern hybridization. Most plants whose progeny segregated in Mendelian ratios for either the NPT II or GUS gene had just one copy of the gene. In this study where both foreign genes were examined in both self and test crosses, no transgenic plant showed Mendelian patterns of inheritance for both foreign traits.