Restriction Site Mapping Research Articles

Identification of a promoter sequence from the BETL1 gene cluster able to confer transfer-cell-specific expression in transgenic maize

The maize (Zea mays L.) betl1 locus, encoding a basal endosperm transfer layer-specific protein, has been mapped and molecularly cloned in its entirety. The locus is shown to consist of three gene copies in the maize inbred line A69Y. To distinguish the three transcription units from the locus name, we have termed them BETL1a, BETL1b, and BETL1c. Two of the copies are expressed, whereas one is inactive and contains retrotransposon-like insertions in both promoter and intron regions. Based on this information, and a restriction site map covering 17 kb around the BETL1 locus, a DNA fragment putatively containing an active promoter sequence was identified. This fragment was tested for its ability to confer transfer-cell-specific expression in transient and stably transformed maize tissues. The transgenic maize plants obtained showed the predicted cell-type specificity of expression restricted to the basal endosperm transfer cells, although there were minor deviations in promoter strength and timing and accumulation of the transgene product from the corresponding BETL-1 endogene expression pattern.

Genetic Diversity in Cotton Assessed by Variation in Ribosomal RNA Genes and AFLP Markers

ABSTRACTVariation in the ribosomal RNA genes (rDNA) and amplified fragment length polymorphism (AFLP) markers has been used to establish the extent of genetic diversity and relatedness in plants. The utility of these methods to detect inter‐ and intra‐specific variation in cotton (Gossypium spp.) has not been reported and could be useful in cultivar identification and in marker assisted selection. The objectives of this study were to: (i) determine the molecular organization of the rDNA genes by restriction enzyme mapping and (ii) assess the level of AFLPs in Old and New World species of cotton. A restriction site map of the rDNA gene structure of G. hirsutum L. cv. TM1 was constructed from DNA digested with 12 restriction enzymes and hybridized to heterologous probes. Four EcoRI‐MseI primer‐pair combinations were used for the AFLP analysis. The rDNA gene structure in cotton was found to be similar to that of most higher plants. The rDNA repeat size was 9.4 kbp in G. hirsutum and G. barbadense L., and 9.6 and 9.8 kbp in G. arboreum L. and G. herbaceum L., respectively. No intraspecific polymorphism was detected in the spacer. The presence of two SspI sites in G. arboreum and G. herbaceum and a single site in G. hirsutum and G. barbadense separated Old and New World cottons. The AFLP method produced a 10‐fold increase in the number of DNA bands per plant, compared with random amplified polymorphic DNA (RAPD) methods. The AFLP data assigned the species–genotypes into groups that corresponded with their origin and/or pedigree relationships.

Chloroplast DNA restriction site variation and phylogeny of the Berberidaceae

Comparative restriction site mapping of the chloroplast genome was performed to examine phylogenetic relationships among 27 species representing 16 genera of the Berberidaceae and two outgroups. Chloroplast genomes of the species included in this study showed no major structural rearrangements (i.e., they are collinear to tobacco cpDNA) except for the extension of the inverted repeat in species of Berberis and Mahonia. Excluding several regions that exhibited severe length variation, a total of 501 phylogenetically informative sites was mapped for ten restriction enzymes. The strict consensus tree of 14 equally parsimonious trees indicated that some berberidaceous genera (Berberis, Mahonia, Diphylleia) are not monophyletic. To explore phylogenetic utility of different parsimony methods phylogenetic trees were generated using Wagner, Dollo, and weighted parsimony for a reduced data set that included 18 species. One of the most significant results was the recognition of the four chromosomal groups, which were strongly supported regardless of the parsimony method used. The most notable difference among the trees produced by the three parsimony methods was the relationships among the four chromosomal groups. The cpDNA trees also strongly supported a close relationship of several generic pairs (e.g., Berberis-Mahonia, Epimedium-Vancouveria, etc.). Maximum likelihood values were computed for the four different tree topologies of the chromosomal groups, two Wagner, one Dollo, and one weighted topology. The results indicate that the weighted tree has the highest likelihood value. The lowest likelihood value was obtained for the Dollo tree, which had the highest bootstrap and decay values. Separate analyses using only the Inverted Repeat (IR) region resulted in a tree that is identical to the weighted tree. Poor resolution and/or support for the relationships among the four chromosomal lineages of the Berberidaceae indicate that they may have radiated from an ancestral stock in a relatively short evolutionary time.

Duplication of the southern white rhinoceros (Ceratotherium simum simum) luteinizing hormone beta subunit gene.

Luteotropic glycoprotein hormones (LGH) include luteinizing hormone (LH) and chorionic gonadotropin (CG). The order Primates is the only phylogenetic clad known to exhibit more than one LGHbeta subunit gene per haploid genome. In the present study, we report the discovery of a second case of LGHbeta gene replication, in the white (w) rhinoceros (r or rhino). The presence of more than one gene was strongly suggested by a complex banding pattern observed on Southern blots of DNA prepared from two unrelated white rhinos. The existence of two LGHbeta genes per haploid genome was estimated by genomic equivalence assay. However, genomic restriction-site mapping studies, together with other findings, suggested that the replicates are probably not tandemly arranged as occurs in primates. A simple band pattern was observed in Southern blots of four other perissodactyl species, indicating that a single-copy LHbeta gene is the consensus condition. Two distinct white rhino LHbeta genomic clones (wrLHbeta1 and wrLHbeta2) were isolated. The nucleotide sequence of wrLHbeta1 was identical with that of wrLHbeta2, except that the latter lacked the consensus mammalian LGHbeta second intron. Sequences of the TATA-containing proximal 5'-flanking regions of the two genes were homologous to at least -57 relative to the site of pituitary transcriptional initiation. We conclude that wrLHbeta1 is the extant form of the ancestral perissodactyl LHbeta gene, whereas wrLHbeta2 is a randomly integrated cDNA element (processed gene) reverse transcribed from a partially spliced ancestral wrLHbeta1 mRNA. That wrLHbeta2 was heritable demonstrates that wrLHbeta1 was transcribed in gametes or early conceptus cells contributing to the germline at some point in time since the divergence of white rhinos from other members of the family Rhinocerotidae. Furthermore, because homologous proximal (pituitary) promoter sequence is present in wrLHbeta2, it can be concluded that the wrLHbeta1 mRNA template from which wrLHbeta2 is derived was transcribed from a secondary promoter located upstream of the consensus TATA-regulated pituitary promoter.

Phylogenetic Reconstruction of the Yeast Genus Kluyveromyces: Restriction Map Analysis of the 5.8S rRNA Gene and the Two Ribosomal Internal Transcribed Spacers

We have constructed restriction site maps of the 5.8S rRNA gene and two ITS regions in 60 strains of Kluyveromyces genus. We test the value of this region as a phylogenetic indicator, and its possible use as a fast and easy method to identify species of this genus. Despite some minor incongruences, our results are in good agreement with previous phylogenetic reconstructions based on the 18S rRNA gene sequencing (Cai et al., 1996; James et al., 1997). A highly significant monophyletic group was formed by K. lactis, K. marxianus, K. aestuarii, K. dobzhanskii and K. wickerhamii, which should be considered the true Kluyveromyces genus. The other species of the genus were grouped with lower bootstrap levels. Finally, the restriction map showed by three K. lactis strains, previously identified as K. marxianus var. drosophilarum, could be interpreted as indicatory of the possible existence of different species.

TheUlmaceae, one family or two? Evidence from chloroplast DNA restriction site mapping

The Ulmaceae is usually split into two subgroups, referred to as either tribes or more commonly subfamilies (Ulmoideae andCeltidoideae). The two groups are separated, with some exceptions, on the basis of leaf venation, fruit type, seed morphology, wood anatomy, palynology, chemistry, and chromosome number. Propositions to separate the two groups as distinct families have never gained general acceptance. Recent morphological and anatomical data have suggested, however, that not only is family status warranted but thatCeltidaceae are more closely related toMoraceae and otherUrticales than toUlmaceae. In order to test these alternative sets of relationships, restriction site mapping of the entire cpDNA was done with nine rare cutting enzymes using 11 genera ofUlmaceae s. l., three other families of theUrticales, and an outgroup family from theHamamelidae. Cladistic analysis of the data indicates thatUlmaceae s. l. is not monophyletic and that distinct families (Ulmaceae andCeltidaceae) are warranted; thatUlmaceae is the sister group toCeltidaceae plus all other families in the order; and thatCannabaceae might be nested withinCeltidaceae. Familial placements of various problematic genera (e.g.Ampelocera, Aphananthe) are resolved and character evolution of key morphological, anatomical, chemical, and chromosomal features are discussed.

Chloroplast DNA restriction site mapping and the phylogeny ofRanunculus (Ranunculaceae)

A chloroplast DNA restriction site map forRanunculus sceleratus (Ranunculaceae) was constructed using 14 restriction endonucleases. The total size of the chloroplast genome is 152.4kb. No inversions were detected relative to the tobacco chloroplast DNA. Cladistic analyses of chloroplast DNA restriction site polymorphism were employed in order to elucidate the phylogeny among 76 species of the genusRanunculus in a wide sense and one species ofTrautvetteria. A total of 341 informative restriction site changes were detected. Parsimony jackknifing, bootstrapping and decay analysis were undertaken in order to evaluate the amount of support for the monophyletic groups. The results suggest that the analysed species ofRanunculus are divisible into two main clades. Only few of the traditional sections and subgenera ofRanunculus are monophyletic. The genusTrautvetteria is nested within a clade comprising, e.g.Ranunculus cymbalaria, R. andersonii, R. lapponicus andR. ficaria. SubgenusBatrachium lies within a larger clade containing, e.g.R. sceleratus andR. hyperboreus. Contractions of the inverted repeat due to parallel deletions of 200–300 bp close to the JSB have occurred in many clades and the phylogenetic distribution of this size reduction was mapped among the species.

Phylogenetic relationships of the macaques (Cercopithecidae: Macaca), as revealed by high resolution restriction site mapping of mitochondrial ribosomal genes.

Molecular phylogenetic relationships among all recognized species within the genus Macaca, were assessed using high-resolution restriction site mapping of the mitochondrial ribosomal genes. By outgroup comparisons to other members of the cercopithecine subfamily, the macaques appear to be a monophyletic assemblage. Within the genus, the relationships are in general consistent with previous genetic studies, though they are less concordant with the separation of the species into four distinct species groups based on modification of the genitalia. Our data support: (1) Macaca sylvanus as sister clade to all Asian macaques; (2) the silenus group as a monophyletic assemblage, with the Sulawesi macaques diverging and colonizing Sulawesi much earlier than previously thought; (3) the fascicularis group as a paraphyletic assemblage, including all non-silenus group Asian macaques; (4) the sinica group as a monophyletic assemblage, possibly derived from a fascicularis-like ancestor; and (5) Macaca arctoides as a separate lineage from the sinica group, also originating from a fascicularis-like ancestor. This study supports the notion that species with more specialized genitalia evolved from less derived taxa, and in general are in agreement with the dispersal scenarios proposed by Fooden (1980) and Delson (1980) for the macaques.

Variation of nuclear ribosomal RNA genes inEragrostis tef(Zucc.) Trotter

Variation in the ribosomal RNA genes (rDNA) was examined to assess the genetic variability among 314 plants representing 28 accessions of Eragrostis tef, an important food crop. A restriction site map was constructed for the species by localization of the BamHI, BglII, DraI, EcoRI, EcoRV, NdeI, SacI, SpeI, XbaI, and XhoI sites. A comparison of this map with those of other grasses showed conservation of sites, especially in the coding region. However, a unique EcoRI site combined with a BamHI site in the 18S region may be of diagnostic value for the species. A BamHI fragment that spans the intergenic spacer was used as an indicator of length variation of rDNA repeat units. rDNA repeat units in E. tef ranged in size from 8.4 to 11.07 kbp. Considerable size variation of rDNA repeats was present among accessions, between individual plants within some accessions, and within single plants. A total of 19 spacer length (sl) phenotypes was observed in 16 accessions in which 11-42 plants were analyzed. A single restriction site polymorphism was detected in PI442115 that was also distinguished by having a single sl variant. Variation in the rRNA genes is a useful indicator of genetic diversity in E. tef germplasm.

Molecular characterization of dematiaceous root endophytes

Sterile dematiaceous fungi are commonly isolated from plant roots. They are often assigned to Mycelium radicis atrovirens , a name originally proposed for black, sterile, fast-growing, pseudomycorrhizal fungi. Dematiaceous fungi isolated from roots may be mutualists, commensalists, or pathogens and, in the absence of sporulation, identification is not possible. Forty-six isolates of dematiaceous fungi from the roots of different hosts and locations were characterized using restriction site mapping of polymerase chain reaction amplified nuclear ribosomal DNA internal transcribed spacers. The restriction site maps were compared to identified dematiaceous mycorrhizal and pseudomycorrhizal fungi. Computer generated trees (UPGMA and parsimony analysis) characterized two unknown isolates as Phialophora finlandia , an ectendomycorrhizal fungus. The majority of the isolates were characterized as Phialocephala fortinii -like. Phialocephala fortinii has been reported as both pathogenic and non-pathogenic in a number of hosts. There was variation within the P. fortinii -like group suggesting intraspecific variation or a species complex.

Chloroplast genome differences between Paspalum dilatatum Poir and the related species P. notatum Flugge

Chloroplast DNA (cpDNA) of Paspalum dilatatum and P. notatum was digested singly or in combination with the restriction endonucleases PstI, PvuII, SalI, KpnI and XhoI. Data obtained from filter hybridization experiments with barley and wheat cpDNA probes were used to construct restriction site maps of the chloroplast genomes of the Paspalum species. The cpDNA fragments were ordered into a circular configuration of approximately 139.3 kbp that contained two inverted repeat regions of approximately 23 kbp and a small and large single-copy region of approximately 11 kbp and 83 kbp, respectively. The cpDNA maps showed that P. dilatatum and P. notatum shared a large number of restriction sites with the proportion of shared restriction sites S=0.90. No restriction site differences were detected in the KpnI maps. Eight species-specific restriction site differences that could be used to identify the cytoplasm of each Paspalum species were identified in the PstI, PvuII, SalI, and XhoI cleavage maps. The overall structural organization of the Paspalum cpDNAs is rather similar to those of most cpDNAs from other plants. The results presented in this study will be of value for exploring further phylogenetic relationships within the genus Paspalum.

A Molecular Phylogeny of Some Bovidae Based on Restriction-Site Mapping of Mitochondrial DNA

Restriction-endonuclease site maps of mitochondrial DNA were constructed for members of the family Bovidae emphasizing species in the Alcelaphinae, Bovinae, and Aepycerotinae. Cladograms weakly supported sister status of the impala, Aepyceros melampus , relative to the Alcelaphinae, and monophyletic status of the Bovini and Tragelaphini. Four members of the Tragelaphini remained an unresolved quadrichotomy, which would be consistent with the inclusion of the eland, Taurotragus oryx , within Tragelaphus . The restriction data are consistent with a radiation for the Tragelaphini at the end of the Miocene.

Mitochondrial DNA restriction site variation within and among five populations of Alaskan coho salmon (Oncorhynchus kisutch)

Mitochondrial DNA (mtDNA) from three Gulf of Alaska and two Bering Sea populations of coho salmon (Oncorhynchus kisutch) was assayed with 21 restriction endonucleases. A restriction site map was constructed for 15 enzymes that recognized hexanucleotide sequences and aligned to the rainbow trout (Oncorhynchus mykiss) gene map using the restriction map and known coho salmon sequences. The restriction site map and gene order of the coho salmon mtDNA genome are consistent with those of rainbow trout. Variation was observed for six enzymes at 10 sites that resulted in eight haplotypes. Variability and divergence observed exceeded those previously observed for allozyme loci in coho salmon. When variability that occurred only in single individuals was eliminated, two haplotypes that differed by four restriction sites remained. Variation at these sites was confirmed from polymerase chain reaction (PCR) amplified fragments. Bering Sea populations exhibited more variation than Gulf of Alaska populations. Explanations for the differences include more recent colonization by or smaller sizes of Gulf of Alaska populations.

Construction of two near-kilobase resolution restriction maps of the 5' regulatory region of the human apolipoprotein B gene by quantitative DNA fiber mapping (QDFM).

Quantitative DNA fiber mapping (QDFM) is a high-resolution technique for physical mapping of DNA. The method is based on hybridization of fluorescently labeled DNA probes to individual DNA molecules stretched on a chemically modified glass surface. We now demonstrate and validate a rapid QDFM-based approach for the mapping of multiple restriction sites and precise localization of restriction fragments in large genomic clones. Restriction fragments of a 70-kb P1 clone (P1-70) containing the 5' region of the human apolipo-protein B gene (APOB) were subcloned and mapped along straightened P1-70 DNA molecules. Multicolor fluorescence in situ hybridization (FISH) and digital image analysis allowed us to rapidly position 29 restriction fragments, ranging in size from 0.5 kb to 8 kb, and to map 43 restriction sites. The restriction map obtained by QDFM was in excellent agreement with information obtained by RecA-assisted restriction endonuclease (RARE) cleavage, long-range PCR, and DNA sequence analyses of the P1-70 clone. These data demonstrate that QDFM is a rapid, reliable method for detailed restriction site-mapping of large DNA clones.

In the public domain

Wonderful as web tools for molecular biology may be, eventually the working scientist must interrupt his or her play with Java Applets, Active-X Buttons, and other such gizmos, and get down to some serious computational molecular biology on his or her own computer. Even here, the web can come in handy, as there are many archives of biological software out there. These act as repositories for ‘public domain’ or ‘shareware’ programs, and also carry reviews or comparisons of commercial offerings. Rather than trying to cover this enormous field in detail, here is a collection of pointers to some of the best resources. Several companies claim to offer ‘comprehensive solutions’ for the molecular biologist's sequence manipulation needs — at a price. Typically, these packages offer restriction-site mapping, contig assembly for sequencing, backwards and forwards translation of DNA sequences, multiple sequence alignment, and much (often, too much) more. Some companies have product information on the web — for example, Geneworks (http://www.ig.com/PRODUCTS/genew_top.html) and PC/GENE (http://www.ig.com/PRODUCTS/pcgene_top.html). There is a good list at http://www.hslib.washington.edu/hsl/bioinfo/software.html. Primer design programs can predict the appropriate temperature for primer annealing and can warn of potential problems with dimerization and hairpin loop formation. Most commercial integrated packages offer primer design, but there are also plenty of free options. There is a good list of primer design programs at the PCR Jump Station (http://www.apollo.co.uk/a/pcr/#pcrs). For die-hard web-users, one of the standard programs, called simply ‘primer’, has been converted into a Java applet by Luca Toldo, so it can be used over the web. It can be found at http://www.embl-heidelberg.de/∼toldo/JaMBW/5/2/index.html. It runs about six-times slower than the normal version, but will work on whatever machine you use — Mac, PC or UNIX — as long as it supports Java. The Department of Genetics of Washington University has a fantastically comprehensive list of programs for the analysis and visualization of phylogenetic relationships (http://evolution.genetics.washington.edu/software.html). It lists more than 80 programs, categorizes them by algorithm and by the computer system on which they run, and provides an excellent evaluation of the strengths and weaknesses of each. Highly recommended. The Protein Databank maintains a list of software (mostly free) for analysing, visualizing, and manipulating proteins structures (http://pdb.pdb.bnl.gov/software.html). A good list of biological software archives is held at Johns Hopkins University (http://www.gdb.org/Dan/softsearch/biol-links.html). Two of the biggest archives it lists are at Indiana University (ftp://ftp.bio.indiana.edu/) and the European Bioinformatics Institute (EBI; http://www.ebi.ac.uk/software/software.html). The software descriptions in the archive at the EBI are fully searchable (http://www.gdb.org/Dan/softsearch/softsearch.html). Bibliographic software is indispensable for the modern biologist. With new packages coming on to the market all the time, the choice can be bewildering, so take a look at the beginner's guide at http://www.chorus.cycor.ca/biblio/biblio1.html. This site also offers up-to-date comparative evaluation of available programs. [email protected] URLs (http://… etc.) can be typed directly into a web browser such as Netscape. A list of the ‘web words’ used in this article can be found in Current Biology 6:494 or its electronic equivalent athttp://BioMedNet.com/ Geneworks PC/GENE List of commercial packages PCR Jump Station ‘Primer’ Java applet Washington University phylogenetics list Protein Databank structural biology list Johns Hopkins University archive list Indiana University software archive European Bioinformatics Institute software archive Searchable software at EBI Beginner's guide to bibliographic software

The Drosophila montium subgroup species. Phylogenetic relationships based on mitochondrial DNA analysis.

Mitochondrial DNA (mtDNA) restriction site maps for three Drosophila montium subgroup species of the melanogaster species group, inhabiting Indian and Afrotropical montium subgroup territories, were established. Taking into account previous mtDNA data concerning six oriental montium species, a phylogeny was established using distance-matrix and parsimony methods. Both genetic diversity and mtDNA size variations were found to be very narrow, suggesting close phylogenetic relationships among all montium species studied. The phylogenetic trees that were constructed revealed three main lineages for the montium subgroup species studied: one consisting of the Afrotropical species Drosophila seguyi, which is placed distantly from the other species, one comprising the north-oriental (Palearctic) species, and one comprising the southwestern (south-oriental, Australasian, Indian, and Afrotropical) species. The combination of the mtDNA data presented here with data from other species belonging to the melanogaster and obscura subgroups revealed two major clusters: melanogaster and obscura. The melanogaster cluster is further divided into two compact lineages, comprising the montium subgroup species and the melanogaster complex species; the species of the other complex of the melanogaster subgroup, yakuba, disperse among the obscura species. The above grouping is in agreement with the mtDNA size variations of the species. Overall, among all subgroups studied, the species of the montium subgroup seem to be the most closely related.

Mitochondrial DNA restriction site map of Cochliomyia macellaria (Diptera:Calliphoridae).

The mitochondrial DNA of the secondary screwworm, Cochliomyia macellaria (F.), was cleaved with 15 restriction endonucleases to produce 54 restriction fragments. Forty-three restriction sites recognized by 12 enzymes were assigned positions on a restriction site map. Both the restriction fragments and the restriction site map were compared with published data from the screwworm, Cochliomyia hominivorax (Coquerel). The restriction site maps were aligned using a combination of highly conserved restriction sites. Estimated nucleotide divergence between the 2 species based on the fragments was 0.052. The divergence based on the restriction site map was 0.093. The data provide a baseline for future population or phylogenetic investigations.

Structure and inheritance of ribosomal DNA variants in cultivated and wild hop, Humulus lupulus L.

Genetic variation was assessed among cultivated and wild hop, Humulus lupulus, by restriction fragment length polymorphisms (RFLPs) of the ribosomal RNA genes (rDNA). Two rDNA length variants of 10.3 and 9.3 kbp represented by three phenotypes designated A, B and C were detected with XhoI. Restriction-site mapping showed that hop rDNA is structurally similar to those of most higher plants. A high level of homogeneity existed in rDNA repeat lengths among the diverse hop genotypes. Generally, phenotype A was predominant in wild and cultivated European and Asian genotypes; phenotype B in North American cultivars; while phenotype C was present only in native North American hop, providing a potential molecular marker for the identification of this germ plasm. The rDNA data provided genetic evidence for the separation of native and cultivated American genotypes and supports the hypothesis that North American hop cultivars are of hybrid origin from European and native American genotypes. The segregation of rDNA phenotypes in four F1 families suggests that a single locus with two co-dominant alleles controls genetic variability for rDNA variants in hop.

Bacillus thuringiensis subsp. aizawai generalized transducing phage 4HD248 : restriction site map and potential for fine-structure chromosomal mapping.

The temperate, generalized transducing phage ΦHD248 from Bacillus thuringiensis subsp. aizawai (H-serotype 7) was shown previously to mediate transduction within subsp. aizawai and between subsp. aizawai and subsp. kurstaki (H-serotype 3a3b). Here it was found to have a broad host range, plating on 7 out of 14 H-serotypes tested. Cotransduction analysis using ΦHD248 revealed two linkage groups in H-serotype 7 : bio-2-his-2-met-1-cit-1-arg-2-lys-3 and trp-9-purC1-thi-1. Preliminary characterization of the ΦHD248 genome revealed a linear double-stranded DNA of 47.15 kb. A restriction site map is reported including a total of 10 sites for the enzymes BamHI, BglII, SalI and SstI Enzyme SalI was found to have a unique cleavage site which if not located in an essential region could be used as a cloning site. The ΦHD248 genome was also shown to lack cohesive ends and appears to have a partial circular permutation.

Ordered yeast artificial chromosome clones representing the Dictyostelium discoideum genome.

High resolution gene maps of the six chromosomes of Dictyostelium discoideum have been generated by a combination of physical mapping techniques. A set of yeast artificial chromosome clones has been ordered into overlapping arrays that cover >98% of the 34-magabase pair genome. Clones were grouped and ordered according to the genes they carried, as determined by hybridization analyses with DNA fragments from several hundred genes. Congruence of the gene order within each arrangement of clones with the gene order determined from whole genome restriction site mapping indicates that a high degree of confidence can be placed on the clone map. This clone-based description of the Dictyostelium chromosomes should be useful for the physical mapping and subcloning of new genes and should facilitate more detailed analyses of this genome. cost of silicon-based construction and in the efficient sample handling afforded by component integration.