Repeating units of xenopus laevis oocyte-type 5S DNA are heterogeneous in length

The restriction enzymes Hind III and Hae III cleave Xenopus laevis 5S DNA at one and three sites, respectively, in each repeating unit of approximately 700 base pairs. The cleavage sites for both enzymes have been located within the repeating unit by denaturation mapping of the restriction fragments. The Hind III products and one of the Hae III fragments are variable in length, indicating heterogeneity in the length of the repeating unit in 5S DNA. This length heterogeneity is confined to the major A + T-rich spacer region. Repeating units differ from each other by discrete quanta of approximately 15 base pairs. The A + T-rich spacer has been shown to consist largely of tandem subrepeats of just this size (Brownlee, Cartwright, and Brown, 1974). We suggest that the repeat-length heterogeneity is due to variable numbers of these subrepeats in the spacer regions of the major repeating units.     

Cleavage map of bacteriophage phiX174 RF DNA by restriction enzymes.

phiX RF DNA was cleaved by restriction enzymes from Haemophilus influenzae Rf (Hinf I) and Haemophilus haemolyticus (Hha. I). Twenty one fragments of approximately 25 to 730 base pairs were produced by Hinf I and seventeen fragments of approximately 40 to 1560 base pairs by Hha I. The order of these fragments has been established by digestion on Haemophilus awgyptius (Hae III) and Arthrobacter luteus (Alu I) endonuclease fragments of phiX RF with Hinf I and Hha1. By this method of reciprocal digestion a detailed cleavage map of phiX RF DNA was constructed, which includes also the previously determined Hind II, Hae III and Alu I cleavage maps of phiX 174 RF DNA (1, 2). Moreover, 28 conditional lethal mutants of bacteriophage phiX174 were placed in this map using the genetic fragment assay (3).     

Cleavage patterns of Drosophila melanogaster satellite DNA by restriction enzymes.

The five satellite DNAs of Drosophila melanogaster have been isolated by the combined use of different equilibrium density gradients and hydrolyzed by seven different restriction enzymes; Hae III, Hind II + Hind III, Hinf, Hpa II, EcoR I and EcoR II. The 1.705 satellite is not hydrolyzed by any of the enzymes tested. Hae III is the only restriction enzyme that cuts the 1.672 and 1.686 satellites. The cleavage products from either of these reactions has a heterogeneous size distribution. Part of the 1.688 satellite is cut by Hae III and by Hinf into three discrete fragments with M.W. that are multiples of 2.3 X 10(5) daltons (approximately 350 base pairs). In addition, two minor bands are detected in the 1.688-Hinf products. The mole ratios of the trimer, dimer and monomer are: 1:6.30 : 63.6 for 1.688-Hae III and 1 : 22.0 : 403 for 1.688-Hinf. Circular mitochondrial DNA (rho = 1.680) is cut into discrete fragments by all of the enzymes tested and molecular weights of these fragments have been determined.     

An approach to a physico-chemical model of olfactory stimulation in vertebrates by single compounds

During recent years, numerous attempts have been made to correlate both quantitative (Davies &; Taylor, 1959; Engen, 1962; Beck, 1964; Engen, Cain &; Rovee, 1968; Cain, 1969; Dravnieks &; Laffoit, 1970; Laffort, 1969a,b) and qualitative (Davies, 1965; Amoore &; Venstrom, 1965; Döving, 1966a,b; Wright &; Michels, 1964; Leveteau &; MacLeod, 1969) odorous properties of single compounds to their molecular properties. These attempts have been only partially successful.In the present paper we will try to explain the several odorous properties of single compounds on the basis of the non-specific properties of odorants involved in solubility.This model is a first approach, and although it gives statistically highly significant relations, it is not as accurate as those advanced with respect to the physical and sensory dimensions of stimuli in the fields of vision and audition.We will first give the present definitions of the most suitable physicochemical parameters, and then advance quantitative and qualitative models for single compounds. Quantitative odorous properties are: odour threshold, rate of change of odour intensity with odorant concentration in the suprathreshold region, and the somewhat controversial upper odour intensity. Qualitative properties refer to odour character.     

The rabbit beta-globin gene contains a large large insert in the coding sequence

We have used the rabbit β-globin DNA plasmid PβG1 (Maniatis et al., 1976) labeled with ³²P as a filter hybridization probe for DNA fragments containing the β-globin gene in restriction endonuclease digests of rabbit liver DNA. The β-globin DNA fragments we detect appear to contain the gene, present in PβG1 DNA, which codes for adult rabbit β-globin. These fragments have been ordered into a physical map of cleavage sites within and neighboring the structural gene in the rabbit genome (Jeffreys and Flavell, 1977). A detailed analysis of β-globin DNA fragments produced by cleavage with restriction endonucleases which are known to cut the β-globin gene has now shown that the β-globin structural gene is not contiguous in rabbit liver DNA, but is interrupted by a 600 base pair DNA segment inserted somewhere within the coding sequence for amino acid residues 101–120 of the 146 residue β-globin chain. Otherwise, the map of cleavage sites within the gene is co-linear with that deduced from the sequence of rabbit β-globin messenger RNA. Preliminary analysis indicates that this insert is also present in the β-globin gene in rabbit brain, kidney, spleen, bone marrow and sperm, and in erythroid cells isolated from the marrow of an anemic rabbit. The insert appears, therefore, to be a general property of the rabbit β-globin gene, even in tissues in which this gene is active, which suggests that the insert is not involved in inactivating the gene in nonerythroid tissues.     

A specific internal RNA polymerase recognition site of VSV RNA is involved in the generation of DI particles.

The 3′ terminal sequences of four different DI particle RNAs ranging in size from 10S to 30S have been determined directly using rapid RNA sequencing methods or deduced, in the case of the fourth DI RNA, from the complementary sequence of a small RNA transcribed from this part of the genome (Schubert et al., 1978). One DI particle (DI 011) contains covalently linked genomic and antigenomic RNA. The 5′ end of this RNA is identical to that of VSV RNA, as determined by annealing for at least 1 kb, as well as to the other DI particle RNAs used in this study. The 3′ ends of the other three DI particle RNAs are exact copies of the common 5′ terminal sequence for 48 nucleotides in two cases and 45 nucleotides in the third. Beyond these complementary regions the sequences are different for each DI RNA. The fact that these regions differ in length by only three nucleotides, despite the wide differences in the overall size of the DI particle RNAs, indicates that if these DIs were formed by the copy-back mechanisms similar to those proposed by Leppert, Kort and Kolakofsky (1977) and Huang (1977), a specific recognition site for the RNA polymerase must be involved in copying the 5′ terminus. We determined the 5′ terminal sequence from position 43–48 at the end of the complementary region and found it to be 5′-GGUCUU-3′. This hexamer is also part of other highly conserved terminal RNA polymerase initiation sites (Keene et al., 1978; Keene, Schubert and Lazzarini, 1979) and may be a specific internal RNA polymerase recognition site. We conclude that this sequence is one of the elements involved in the genesis of DI particle chromosomes containing short complementary sequences at their termini. The ability of the polymerase to resume synthesis at or near a specific recognition site is discussed.     

Multiple,heterogeneous actin genes in Dictyostelium

We have used an actin gene-containing restriction fragment of plasmid M6 (Kindle and Firtel, 1978) to select a second actin gene-containing plasmid which we have named pDd actin 2. This plasmid has been shown to contain two actin genes separated by 350 bp of nonactin DNA. When heteroduplexes are formed between any two of the three actin genes present in chimeric plasmids, the region of homology is 1100 ± 100 bp. This is close to the minimum length required to code for actin protein. The 1100 bp region of intergene homology corresponds to the 1100 bp homology observed between M6 and the two actin cDNA plasmids pcDd actin B1 and pcDd actin A1 (Bender et al., 1978). We have no evidence for additional sequences common to either the 3′ or 5′ ends of the 1100 ± 100 bp region of intergene homology. Thermal denaturation experiments show that different pairs of actin genes are diverged from each other by as much as 6–8%. There are two size classes of mRNA complementary to the three actin genes. These have lengths of 1.25 and 1.35 kb as determined on methyl mercuric hydroxide-containing agarose gels. The possible linkage of these three actin genes to other actin genes is discussed.     

RNA sequences specifically associated with mouse intracisternal A particles.

Electron microscopic examination of the histone H1-depleted, folded genomes of Drosophila melanogaster reveals that they are composed of long cylindrical cables of about 100 Å diameter. Limited single-strand nicking with DNAase I relaxes the 100 Å fibers to a “beads-on-a-string” structure, showing the nucleosomes and internucleosome DNA.Based on these results and other available data, we have constructed a detailed space-filling model for the higher order DNA coiling in chromatin, starting with the symmetrical nucleosome core previously described (Weintraub, Worcel and Alberts, 1976). The model defines the path of the DNA helix and the nucleosome arrangement along the DNA coil for both the 100 Å and the 200–300 Å fibers.Following Sobell et al. (1976), we believe that the DNA is coiled in the 100 Å nucleofilament in a uniform left-handed supercoil of about 90 base pairs (bp) per turn and 47 Å pitch; the 140 bp symmetrical nucleosome cores align themselves along this uniform DNA superhelix so that the isologous outer surfaces of adjacent nucleosomes touch and the internucleosome spacer DNA coils between them. A few single-strand discontinuities [about one nick per 85 kilobases (kb); Benyajati and Worcel, 1976] in the H1-depleted 100 Å fiber can thus relax the negatively supercoiled internucleosome DNA generating the “beads-on -a-string” appearance.We propose that histone H1 binds to the 100 Å diameter superhelix and coils it into tightly packed, 110 Å pitch super-superhelices (“solenoids;” Finch and Klug, 1976) of variable diameter (between 200–300 Å). In our model, the “thick” 200–300 Å fiber is stabilized at metaphase by histone H1-H1 heterologous interactions between adjacent helical turns of the nucleofilament, and the internucleosome spacer DNA is located on the outside. Symmetry considerations demand that changes in the length of the repeat should lead to variations in the number of nucleosomes per helical turn and in the handedness of these turns in the 200–300 Å metaphase fiber.     

Banding patterns of the chromosomes of the Rhesus monkey (Macaca mulatta)

The Rhesus monkey (Macaca mulatta) has 21 pairs of chromosomes, many of which can easily be confused with one another by the traditional staining methods. By using the method of banding with trypsin (Seabright, 1971) we have been able to characterize the various pairs of homologous chromosomes and we have classified them by following the criteria adopted by Rothfels & Siminovich (1958). The evolutionary meaning of the results are discussed.     

Replication of polyoma DNA in isolated nuclei. VII. Initiator RNA synthesis during nucleotide depletion.

Earlier experiments demonstrated that the Okazaki fragments synthesized during discontinuous polyoma DNA synthesis in isolated nuclei at their 5′ ends contained structural elements consisting of polyribonucleotides starting with ATP or GTP (Reichard et al., 1974). These structures could be released by digestion with pancreatic DNAase and were named initiator RNA. They consist of a large family of polyribonucleotides differing in base sequence but having a common size of about a decanucleotide. We now demonstrate that limitation of DNA synthesis by low concentrations of deoxyribonucleoside triphosphates in parallel limits the synthesis of initiator RNA. This is additional evidence for the primer function of initiator RNA. When ribonucleoside triphosphates other than ATP were deleted from the incubation medium only a small decrease of DNA and initiator RNA synthesis occurred. Under those conditions deoxyribonucleotides substituted for ribonucleotides and were incorporated internally into the primer. From this result as well as the insensitivity of initiator RNA synthesis to α-amanitin (Reichard &; Eliasson, 1979) we suggest that a mammalian counterpart to primase, the dnaG gene product of Escherichia coli(Rowen &; Kornberg, 1978a), catalyzes the synthesis of initiator RNA.     

Kinship and covariance

Price's (1970) covariance theorem can be used to derive an expression for gene frequency change in kin selection models in which the fitness effect of an act is independent of the genotype of the recipient. This expression defines a coefficient of relatedness which subsumes r(Wright, 1922), b(Hamilton, 1972), ρ (Orlove &; Wood, 1978), and R(Michod &; Hamilton, 1980). The new coefficient extends the domain of Hamilton's rule to models in which the average gene frequency of actors differs from that of recipients.     

The role of migration in the genetic structure of populations in temporally and spatially varying environments II. Island Models

The Island Model introduced by Sewall Wright (1951) has proven to be a useful construction for studying the interaction of genetic drift, population subdivision, and mutation. Interest in the model has recently increased because of its relevance to certain questions involving the rate of differentiation of sub-populations under the neutral allele hypothesis (e.g., Smith, 1970; Latter, 1973). It is perhaps the only realistic population structure in which the test for neutrality proposed by Lewontin and Krakauer (1973) is valid (Lewontin and Krakauer, 1975). If data from natural populations is to be compared to the predictions of the Island Model, it is desirable to have an alternative model with the same migration pattern but with natural selection operating. In this paper one such model will be introduced where the stochastic element comes from random fluctuations in the environment rather than from genetic drift. The model is a direct extension of the one in the previous paper in this series (Gillespie, 1975) which dealt with a population which is subdivided into two patches with restricted migration between them.     

Replication of a mammalian genome: the role of de novo protein biosynthesis during S phase

ts14 is a temperature-sensitive Chinese hamster lung cell mutant that ceases protein biosynthesis within a short time of transfer to nonpermissive temperature (Haralson and Roufa, 1975; Roufa and Haralson, 1975; Roufa and Reed, 1975). This mutant contains a revertible, presumably a point mutation that renders its 60S ribosomal subunit thermolabile (Haralson and Roufa, 1975). In this report, we describe the relationship between the conditional ability of ts14 to synthesize protein during S phase and the replication of its DNA.After transfer to nonpermissive temperature (39°C), where ts14 synthesizes protein at a rate approximately 20 fold less than wild-type cells, synchronous cultures of the mutant performed all the processes required for replication of their DNA. During prolonged incubations at nonpermissive temperature, S phase ts14 completed approximately one round of DNA replication semi-conservatively as judged by density-transfer experiments. Pulse-labeling experiments performed on S phase cells revealed that ts14 synthesized the intermediates of discontinuous DNA replication at nonpermissive and permissive temperatures at similar rates. In these tests, the mutant was not substantially different from wild-type at both culture temperatures. At the nonpermissive temperature, however, ts14 synthesized significantly less nuclear protein (that is, histone) than did wild-type cells, and the mutant's chromatin appeared deficient in histone by virtue of its increased sensitivity to nuclease.     

Long range periodicities in mouse satellite DNA.

Escherichia coli restriction enzyme II breaks mouse satellite DNA into fragments which form a series of bands on gel electrophoresis. The DNA in the strongest band has a length of 220 to 260 nucleotide pairs and the other bands are multiples of this length. It is shown that these fragments are linked together in long arrays in the satellite sequence. The reassociation register of the DNA is about half the length of the 220 to 260 nucleotide pair fragment. In the electrophoresis pattern of the Eco RII² fragments other weaker bands can be seen. The stronger bands of the minor patterns fall half-way between the bands of the main pattern and the smallest is 120 to 130 nucleotide pairs long. The properties of the minor fragments suggest short spacings of the restriction site which have been produced by unequal crossing-over. The extents of divergence and unequal crossing-over are estimated. From this analysis and the sequence analysis described in the accompanying paper (Biro et al., 1975) it is proposed that mouse satellite DNA consists of an hierarchy of four periodicities which reflect stages in the evolution of the sequence.Digestion of mouse satellite DNA with Hae III produces fragments with the same sizes as those produced by Eco RII, but the yields are much lower. It is suggested that Hae III sites have been introduced by divergence and subsequently spread by unequal crossing-over.     

A general theory of chromosome pairing based on the palindromic DNA model of Sobell with modifications and amplifications

A model for chromosome pairing is presented. It is based on the presence of segments of symmetric sequences of bases (palindromes) in the DNA at specific places in the chromosome. Palindromic DNA has been characterized by Wilson &; Thomas (1974), who state that these sequences are a regular feature of eukaryotic DNA. Sobell (1972) has suggested that they may be involved in synapsis and genetic recombination. Sobell's model is modified and amplified in an attempt to develop a general theory of chromosome pairing that explains congressional pairing, synapsis, non-homologous pairing, the initiation of crossing over, and interference.     

The fine structure of Ameson pulvis (Microspora,Microsporida) and its implications regarding classification and chromosome cycle

Nosema pulvisPerez, 1905, Ameson pulvis (Perez) Sprague, 1977, in muscles of the crabs Carcinus maenas and C. mediterraneus from the coast of France, was observed with the electron microscope. It was found to be structurally similar to the type species A. michaelis (Sprague, 1970). Sprague, 1977, having moniliform sporogonial plasmodia, unikaryotic sporoblasts, and hirsute sporulation stages. It is treated as distinct from A. michaelis because it has slightly smaller spores (by comparison with syntype material of A. michaelis) and appears to have fewer coils in the polar filament. The results require the removal of the genus Ameson from the family Nosematidae Labbé, 1899, where Sprague (1977) had placed it under the erroneous supposition that its sporoblasts are diplokaryotic. Ameson is transferred to family Unikaryonidae Sprague, 1977. Ameson is distinguished from PereziaLéger and Duboscq, 1909, shown by Ormieres et al. to have a similar developmental pattern, by presence of appendages on its sporulation stage. A. nelsoni (Sprague, 1950), the third, and only other species of Ameson, lacks the appendages and is transferred to genus Perezia.     

A note on the sampling theory for infinite alleles and infinite sites models

This note considers sampling theory for a selectively neutral locus where it is supposed that the data provide nucleotide sequences for the genes sampled. It thus anticipates that technical advances will soon provide data of this form in volume approaching that currently obtained from electrophoresis. The assumption made on the nature of the data will require us to use, in the terminology ofKimura (Theor. Pop. Biol.2, 174–208 (1971)), the “infinite sites” model of Karlin and McGregor (Proc. Fifth Berkeley Symp. Math. Statist. Prob.4, 415–438 (1967)) rather that the “infinite alleles” model of Kimura and Crow (Genetics49, 174–738 (1964)). We emphasize that these two models refer not to two different real-world circumstances, but rather to two different assumptions concerning our capacity to investigate the real world. We compare our results where appropriate with corresponding sampling theory of Ewens (Theor. Pop. Biol.3, 87–112 (1972)) for the “infinite alleles” model. Note finally that some of our results depend on an assumption of independence of behavior at individual sites; a parallel paper byWatterson (submitted for publication (1974)) assumes no recombination between sites. Real-world behavior will lie between these two assumptions, closer to the situation assumed by Watterson than in this note. Our analysis provides upper bounds for increased efficiency in using complete nucleotide sequences.     

Isolation and some properties of DNA coding for tRNA1met from Xenopus laevis

DNA containing the reiterated genes for tRNA₁^met has been partially purified from Xenopus laevis by centrifugation in actinomycin C₁-CsCl and Ag⁺-Cs₂SO₄ gradients. These gradients separate the tRNA₁^met genes from those coding for tRNA₂^met and tRNA^val, thus confirming our earlier suggestion that these genes are not intermingled with each other (Clarkson, Birnstiel, and Purdom, 1973). The gradients also demonstrate the existence of a minor 5S DNA fraction which appears to differ from that previously isolated by Brown, Wensink, and Jordan (1971).When the enriched tDNA₁^met is digested to completion with either of the restriction endonucleases EcoRI or Hpa I, the tRNA₁^met genes are predominantly found within DNA fragments that are about 3100 base pairs long. A partial digestion with EcoRI shows that these fragments arise from the regular spacing of the enzyme restriction sites. The 3100 base pair EcoRI fragments are cleaved by Hpa I into fragments of two size classes, one of which is about 2200 base pairs long and contains the tRNA₁^met genes. The shorter fragments are about 700 base pairs long, and they appear to contain genes coding for at least one other kind of tRNA species. X. laevis tDNA₁^met thus comprises tandemly repeated DNA whose component parts show little if any length heterogeneity.