Co-dominant markers for the selection of somatic dihybrids and polyhybrids

Resistance to ouabain and puromycin are shown to represent very useful co-dominant characters for the selection of somatic hybrids between mammalian cells, after fusion with polyethylene glycol. We therefore used, with success, a number of Chinese hamster and mouse cell lines carrying these markers in association with thymidine kinase and hypoxanthine-guanine-phosphoribosyl transferase deficiency for selection of hybrids of triparental origin.     

Zum Problem der Genetischen Rekombination von Bakteriophagen

Summary Four equivalent triparental-3 factor-3 alleles crosses of the type 121×212×322 are performed with linked markers in bacteriophage T1. Total recombinants and specially the biparental doubles (BD) of the type 111 and the triparental doubles (TD) of the type 311 are scored. The results support the idea of a topography and give therefore no conclusive evidence whether the process of cooperation is restricted to two parental structures or not.

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The occurrence of three allelic markers in one particle of phage T4

Summary The analysis of the progeny from three-allelic, triparental one-factor crosses in T4 showed a fraction of phages carrying allthree markers inone particle. Thesethree-allelic heterozygotes were observed in therII-region as well as in the host range gene. Their frequency (0.01–0.04% of total yield) was in the range expected from theory. This result is discussed with respect to the current picture of circular permutation in phage T4.With 1 Figure in the Text     

Genetic mapping: Chromosomes 2–5

Summary Genetic maps of chromosomes 2 and 4 constructed from pair-wise lod score data from family studies and regional assignments for markers are presented. Two loci are mapped on chromosome 2 and multiple crossing-over is suggested as an explanation for the poor fit to the data in females. The best map of chromosome 4 gives the genetic locations of five markers with the Stoltzfus (SF) blood group distal to MNS on the long arm and GC close to the centromere on the short arm. This position for GC is outside its provisional regional assignment and possible reasons for this discrepancy are discussed. The GM-PI linkage group has a score of less than-1.0 with chromosome 4 suggesting that it may be excluded from that chromosome.The regional assignment for markers on chromosome 2–5 are also shown.     

Electron microscopic evidence for the axial rotation and inter-domain flexibility of the Fab regions of immunoglobulin G

The Fab arms of immunoglobulin G (IgG) have long been known to hinge about their joint with the Fc subunit. Using monoclonal antibodies bound to influenza haemagglutinin (HA) as position markers, we now show that these arms can also rotate about their long axis with respect to Fc. We also show that when two IgGs are bound cyclically with two HA molecules, the arms can bend between the variable and constant domains to accommodate bond angle constraint.     

LuxAB标记荧光假单胞菌的菜心根际定殖研究

目的:利用三亲本杂交方法将luxAB发光酶基因标记至荧光假单胞菌PF20001上,所获得的标记菌株PF20001-Lux能稳定发光。将该标记菌株制成微生物制剂,接种菜心进一步研究它在菜心根际的定殖动态和散布规律。方法:利用三亲本杂交方法将luxAB发光酶基因标记至荧光假单胞菌PF20001上,将该菌施与菜心生长土壤中,通过接合子发光检测及发光菌落的平板计数来分析荧光假单胞菌在菜心根际的定殖分布情况。结果:PF20001-Lux在根系周围的土壤中的有一定的定殖率,在根内主要定殖在3～4cm根内。PF20001-Lux在菜心种植后7d前就达到最高定殖水平达3.8×105CFU/g,随后逐渐下降。结论:PF20001-lux在菜心根际具备良好的适应能力。     

Transposon-induced mutants of the cyanobacterium Anabaena sp. PCC7120 capable of ammonia liberation

Summary A triparental conjugation technique (using pRL10630 plasmid) was used in creation and characterization of three transposon-induced mutants of Anabaena PCC7120 which are capable of extracellular ammonia liberation in the absence and /or presence of a glutamate analogue (MSX). Results suggest that such mutants can potentially serve as suitable biofertilizer to support crops without addition of the glutamate analogue.     

A map of the distal region of the long arm of human chromosome 21 constructed by radiation hybrid mapping and pulsed-field gel electrophoresis

We have used radiation hybrid (RH) mapping and pulsed-field gel electrophoresis (PFGE) to determine the order and positions of 28 DNA markers from the distal region of the long arm of human chromosome 21. The maps generated by these two methods are in good agreement. This study, combined with that of D. R. Cox et al. (1990, Science 250:245-250), results in an RH map that covers the long arm of chromosome 21 (21q). We have used a subtelomeric probe to show that our map includes the telomere and have identified single-copy genes and markers within 200 kbp of the telomere. Comparison of the physical and RH maps with genetic linkage maps shows "hot spots" of meiotic recombination in the distal region, one of which is close to the telomere, in agreement with previous cytogenetic observations of increased recombination frequency near telomeres.     

Cross-species transfer of nuclear microsatellite markers: potential and limitations

Molecular ecologists increasingly require 'universal' genetic markers that can easily be transferred between species. The distribution of cross-species transferability of nuclear microsatellite loci is highly uneven across taxa, being greater in animals and highly variable in flowering plants. The potential for successful cross-species transfer appears highest in species with long generation times, mixed or outcrossing breeding systems, and where genome size in the target species is small compared to the source. We discuss the implications of these findings and close with an outlook on potential alternative sources of cross-species transferable markers.     

Long tomato microsatellites are predominantly associated with centromeric regions.

Microsatellites as genetic markers are used in many crop plants. Major criteria for their usability as molecular markers include that they are highly polymorphic and evenly spread throughout a genome. In tomato, it has been reported that long arrays of tetranucleotide microsatellites containing the motif GATA are highly clustered around the centromeres of all chromosomes. In this study, we have isolated tomato microsatellites containing long arrays (> 20 repeats) of the dinucleotide motifs GA, GT, AT, as well as GATA, assessed their variability within Lycopersicon esculentum varieties and mapped them onto a genetic map of tomato. The investigated microsatellite markers exhibited between 1 and 5 alleles in a diverse set of L. esculentum lines. Mapping of the microsatellites onto the genetic map of tomato demonstrates that, as previously shown, GATA microsatellites are highly clustered in the regions of the tomato centromeres. Interestingly, the same centromeric location was now found for long dinucleotide microsatellite markers. Because of this uneven distribution, genetic mapping of the entire tomato genome using long dinucleotide microsatellites will be very difficult to achieve and microsatellite markers with shorter arrays of microsatellites could be more suitable for mapping experiments albeit their lower level of polymorphism. Some microsatellite markers described in this study might provide a useful tool to study the molecular structure of tomato centromeric regions and for variety identification.     

Genomic targeting and high-resolution mapping of the domestication gene Q in wheat.

The Q locus is largely responsible for the domestication of bread wheat. Q confers the free-threshing character of the spike and influences other important agronomic traits. Using chromosome deletion lines, Q was placed on the physical map within a submicroscopic segment of the long arm of chromosome 5A. We targeted markers to the segment by comparative mapping of anonymous RFLP clones, AFLP, and mRNA differential display analysis of deletion lines 5AL-7 and -23, which have deletion breakpoints that flank the Q locus. Differentially expressed sequences detected fragments at various loci on group 5 chromosomes suggesting that Q may be a regulatory gene. We identified 18 markers within the Q gene deletion interval and used them to construct a genetic linkage map of the region in F2 populations derived from chromosome 5A disomic substitution lines. The genetic map corresponding to the deletion segment was 20-cM long, and we identified markers as close as 0.7 cM to the Q gene. An estimate of base pairs per centimorgan within the region is 250 kb/cM, an 18-fold increase in recombination compared with the genomic average. Genomic targeting and high-density mapping provide a basis for the map-based cloning of the Q gene.     

Genetic clues to the origin of the apple

Molecular genetic markers complement archaeological, breeding and geographical investigations of the origins, history and domestication of plants. With increasing access to wild apples from Central Asia, along with the use of molecular genetic markers capable of distinguishing between species, and explicit methods of phylogeny reconstruction, it is now possible to test hypotheses about the origin of the domesticated apple. Analyses of nuclear rDNA and chloroplast DNA (cpDNA) sequences indicate that the domesticated apple is most closely related to series Malus species. Moreover, the occurrence of a shared 18-bp duplication in the cpDNAs of wild and cultivated apple supports the close relationship between them. Hypotheses about the hybridization and the origin of the domesticated apple cannot be rejected completely until more variable, phylogenetically informative markers are found.     

Zebrafish Genetic Map with 2000 Microsatellite Markers

The zebrafish is the first vertebrate organism used for large-scale genetic screens seeking genes critical to development. These screens have been quite successful, with more than 1800 recessive mutations discovered that speak to morphogenesis of the vertebrate embryo. The cloning of the mutant genes depends on a dense genetic map. The 2000 markers we present here, using microsatellite (CA) repeats, provides 1.2-cM average resolution. One centimorgan in zebrafish is about 0. 74 megabase, so, for many mutations, these markers are close enough to begin positional cloning by YAC walks.     

Genetic heterogeneity in X-linked amelogenesis imperfecta.

The AMELX gene located at Xp22.1-p22.3 encodes for the enamel protein amelogenin and has been implicated as the gene responsible for the inherited dental abnormality X-linked amelogenesis imperfecta (XAI). Three families with XAI have been investigated using polymorphic DNA markers flanking the position of AMELX. Using two-point linkage analysis, linkage was established between XAI and several of these markers in two families, with a combined lod score of 6.05 for DXS16 at theta = 0.04. This supports the involvement of AMELX, located close to DXS16, in the XAI disease process (AIH1) in those families. Using multipoint linkage analysis, the combined maximum lod score for these two families was 7.30 for a location of AIH1 at 2 cM distal to DXS16. The support interval around this location extended about 8 cM proximal to DXS92, and the AIH1 location could not be precisely defined by multipoint mapping. Study of recombination events indicated that AIH1 lies in the interval between DXS143 and DXS85. There was significant evidence against linkage to this region in the third family, indicating locus heterogeneity in XAI. Further analysis with markers on the long arm of the X chromosome showed evidence of linkage to DXS144E and F9 with no recombination with either of these markers. Two-point analysis gave a peak lod score at DXS144E with a maximum lod score of 2.83 at theta = 0, with a peak lod score in multipoint linkage analysis of 2.84 at theta = 0. The support interval extended 9 cM proximal to DXS144E and 14 cM distal to F9.(ABSTRACT TRUNCATED AT 250 WORDS)     

Molecular mapping of two dwarfing genes differing in their GA response on chromosome 2H of barley

The two recessive dwarfing mutants gai (GA-ins) and gal (GA-less), differing in their response to exogenously applied gibberellic acid (GA₃), were mapped in the centromere region and on the long arm, respectively, of the barley chromosome 2H. The gene gai, which determines reduced plant height and GA insensitivity pleiotropically, was found to co-segregate with the two RFLP markers Xmwg2058 and Xmwg2287. Both markers are known to map close to the centromere. The GA-sensitive dwarfing gene gal was found to be linked to the three co-segregating RFLP markers Xmwg581, Xmwg882 and Xmwg2212 (proximal) and XksuG5 (distal) by 3.6 and 9.5. cM, respectively. The distance between the two mutant loci was estimated to be about 55 cM. Homoeologous relationships between the dwarfing genes within the Triticeae are discussed. Received: 11 December 1998 / Accepted: 11 February 1999     

A linkage map of three anonymous human DNA fragments and SOD-1 on chromosome 21

Using DNA polymorphisms adjacent to single-copy genomic fragments derived from human chromosome 21, we initiated the construction of a linkage map of human chromosome 21. The probes were genomic EcoRI fragments pW228C, pW236B, pW231C and a portion of the superoxide dismutase gene (SOD-1). DNA polymorphisms adjacent to each of the probes were used as markers in informative families to perform classical linkage analysis. No crossing-over was observed between the polymorphic sites adjacent to genomic fragments pW228C and pW236B in 31 chances for recombination. Therefore, these fragments are closely linked to one another (theta = 0.00, lod score = 6.91, 95% confidence limits = 0-10 cM) and can be treated as one 'locus' with four high-frequency markers. There is a high degree of non-random association of markers adjacent to each of these two probes which suggests that they are physically very close to one another in the genome. The pW228C - pW236B 'locus' was also linked to the SOD-1 gene (theta = 0.07, lod score = 4.33, 95% confidence limits = 1-20 cM). On the other hand, no evidence for linkage was found between the pW228C-pW236B 'locus' and the genomic fragment pW231C (theta = 0.5, lod score = 0.00). Based on the fact that pW231C maps to 21q22.3 and SOD-1 to 21q22.1, we suggest that the pW228C-pW236B 'locus' lies in the proximal long arm of chromosome 21. These data provide the outline of a linkage map for the long arm of chromosome 21, and indicate that the pW228C-pW236B 'locus' is a useful marker system to differentiate various chromosome 21s in a population.     

The identification of random amplified polymorphic DNA markers for daylength insensitivity in oat.

Daylength insensitive accessions of Avena sativa L. are being used to develop cultivars that will flower normally when grown under short or long photoperiods. Field data indicate that the insensitivity trait is under the control of a single dominant gene, designated Di1. The random amplified polymorphic DNA (RAPD) technique and bulk segregant analysis of daylength sensitive and insensitive plants were used to find markers for this gene. Five of 200 random decamer primers tested produced polymorphic bands, which were shown to be linked to the trait using 30 homozygous insensitive and 30 homozygous sensitive F3 individuals. Three of the markers produced a band in the presence of the dominant allele, and two in its absence. Segregation analysis showed that markers 221 and 136 could be mapped to within 9.8 +/- 4.6 and 13.9 +/- 5.4 cM of the trait, respectively; that is, close enough to be useful in a breeding program. A study of different cultivars suggested that the band produced by primer 136 is actually the more closely linked marker and the only one present in the original Di1 gene donor CAV2700. The possibility of using both markers in populations derived from different cultivars is discussed.     

Retardation of bone development in the Koshima troop of Japanese macaques

Retardation of bone development was observed in the Koshima troop of free ranging Japanese macaques. In the control group, epiphyseal unions of appendicular long bones generally started to close at about 4 yrs of age and were completed at about 8 or 9 yrs of age. Limb bone unions of the Koshima troop, however, started to close at about 9 yrs of age and completely closed at about 15 yrs of age. In the epiphyseal unions of trunk and girdle bones, the Koshima troop again showed a retardation of closure compared with the control group. Until long bones reached their full length, that is, until about 15 yrs of age, their size was small in the Koshima troop compared with the control group, though the sample size of the Koshima troop was small. After 15 yrs of age, however, many osteometrical measurements of the Koshima troop were nearly the same as controls. A prolonged growing duration compensated for the slow growth and allowed them to become as large as controls. This prolongation may be an adaptation in response to small size during the developmental period. In some parts of the body, however, Koshima macaques failed to reach the adult size of controls. Males were less likely than females to reach full size. Causes of the retardation and small size in the Koshima troop are discussed, but they remain open to further studies.     

Comparison of the physical and recombination maps of the mouse X chromosome

The locations of five random mouse genomic DNA markers and five cloned genes, including the genes for clotting factors VIII and IX (Cf-8 and Cf-9), Duchenne muscular dystrophy (Dmd), phosphoglycerate kinase-1 (Pgk-1), and alpha-galactosidase (Ags), on the mouse X chromosome were determined by in situ hybridization. The five random DNA markers provide new genetic loci with useful restriction fragment length polymorphisms between mouse strains and species, including one locus close to the centromeric region of the mouse X chromosome. The physical map and the recombination map of these loci on the X chromosome were compared. There was good agreement in the order of loci. Relative distances between loci were consistent along the X chromosome, with the exception of the telomeric end of the long arm, where the recombination fraction observed between loci closely associated on the physical map was higher than that between similarly spaced markers located in the proximal region of the X chromosome. These results are discussed in comparison to the human X-chromosome map.     

Genome-scale patterns in the loss of heterozygosity incidence in Saccharomyces cerevisiae

Former studies have established that loss of heterozygosity can be a key driver of sequence evolution in unicellular eukaryotes and tissues of metazoans. However, little is known about whether the distribution of loss of heterozygosity events is largely random or forms discernible patterns across genomes. To initiate our experiments, we introduced selectable markers to both arms of all chromosomes of the budding yeast. Subsequent extensive assays, repeated over several genetic backgrounds and environments, provided a wealth of information on the genetic and environmental determinants of loss of heterozygosity. Three findings stand out. First, the number of loss of heterozygosity events per unit time was more than 25 times higher for growing than starving cells. Second, loss of heterozygosity was most frequent when regions of homology around a recombination site were identical, about a half-% sequence divergence was sufficient to reduce its incidence. Finally, the density of loss of heterozygosity events was highly dependent on the genome’s physical architecture. It was several-fold higher on short chromosomal arms than on long ones. Comparably large differences were seen within a single arm where regions close to a centromere were visibly less affected than regions close, though usually not strictly adjacent, to a telomere. We suggest that the observed uneven distribution of loss of heterozygosity events could have been caused not only by an uneven density of initial DNA damages. Location-depended differences in the mode of DNA repair, or its effect on fitness, were likely to operate as well.