Construction and characterization of a YAC library with a lowfrequency of chimeric clones from flow-sorted human chromosome 9

Human chromosome 9 DNA, flow-sorted from somaticcell hybrid PK-87-9, has been used to construct two complete digest YAC libraries. The combined representation of chromosome 9 in these libraries, estimated by hybridization of chromosome 9-specific sequences to YAC colony grids, is 95%. The frequency of chimeric clones, analyzed by fluorescence in situ hybridization of chromosome 9 YACs to human metaphase chromosomes, was estimated to be 4%. These libraries provide a resource for physical mapping and for moving from genetic markers to disease loci on chromosome 9.     

3D 13C/1H NMR-based assignments for side-chain resonances of Lactobacillus casei dihydrofolate reductase. Evidence for similarities between the solution and crystal structures of the enzyme

Summary ¹³C-based three-dimensional ¹H–¹H correlation experiments have been used to determine essentially complete ¹³C and ¹H resonance assignments for the amino acid side chains of uniformly ¹³C/¹⁵N labelled L. casei dihydrofolate reductase in a complex with the drug methotrexate. Excellent agreement is observed between these assignments and an earlier set of partial assignments made on the basis of correlating nuclear Overhauser effect and crystal structure data, indicating that the tertiary structure of the enzyme is similar in solution and in the crystal state.To whom correspondence should be addressed.     

A 610 kb YAC clone harbors 7 cM of tomato (Lycopersicon esculentum) DNA that includes themale sterile 14 gene and a hotspot for recombination

Pollen development requires both sporophytic and gametophytic gene expression. We are using a map-based cloning technique to isolate sporophytic genes which, when mutant, cause pollen abortion and a male sterile (ms) phenotype in tomato (Lycopersicon esculentum). We have genetically characterized onems locus (ms14) using RFLP analysis and identified flanking markers. High-resolution genomic physical mapping indicates that thems14 locus is located in a ～300 kb region. We have identified a YAC clone with an insert size of ～610 kb that contains thems14-linked markers, reflects the organization of the physical map and therefore most probably contains thems14 gene. In addition, we present evidence that the relationship between physical and genetic distance in this chromosomal region changes abruptly from ～105–140 kb/cM to less than 24 kb/cM, and suggest that the TG393-TG104 region is a hotspot for recombination.     

Assembly of the cell division protein FtsZ into ladder-like structures in the aerial hyphae of Streptomyces coelicolor

In the filamentous bacterium Streptomyces coelicolor , the cell division protein FtsZ is required for the conversion of multinucleoidal aerial hyphae into chains of uninucleoidal spores, although it is not essential for viability. Using immunofluorescence microscopy, we have shown that FtsZ assembles into long, regularly spaced, ladder-like arrays in developing aerial hyphae, with an average spacing of about 1.3 μm. Within individual hyphae, ladder formation was relatively synchronous and extended for distances over 100 μm. These ladders were present only transiently, decreasing in intensity as chromosomes separated into distinct nucleoids and disappearing upon the completion of septum formation. Evidence from the overall intensity of immunofluorescence staining suggested that ladder formation was regulated in part at the level of the accumulation and degradation of FtsZ within individual aerial hyphae. Finally, FtsZ ladder formation was under developmental control in that long arrays of FtsZ rings could not be detected in certain so-called white mutants ( whiG , whiH and whiB ), which are blocked in spore formation. The assembly of FtsZ into ladders represents the earliest known molecular manifestation of the process of spore formation, and its discovery provides insight into the role of whi genes in the conversion of aerial hyphae into chains of spores. We have also described a novel use of a cell wall-staining technique to visualize apical tip growth in vegetatively growing hyphae.