Gaucher disease (GD) type 1 is the most prevalent lysosomal storage disease and the most prevalent genetic disease among the Ashkenazi Jews. The defective activity of acid /3-glucosidase is the enzymatic basis of GD and is inherited as an autosomal recessive trait. To investigate the genetic basis of Ashkenazi Jewish GD type 1, a cDNA encoding acid /3-glucosidase was isolated from a cDNA library constructed using splenic poly(A)+RNA from a patient. The cDNA was sequenced to identify mutations, and the presence of a single missense mutation in the patients' genome was confirmed by selective oligonucleotide hybridization and by restriction endonuclease digestion analyses of amplified genomic sequences. This G-A transition (Arg-119 to Gln-119) was present heterozygously in the index patient and his affected third cousin but was not present in normal non-Jewish individuals. This mutation is the second single base mutation found in Ashkenazi Jewish GD type 1 patients. Furthermore, results obtained with the affected third cousin suggest that at least three mutant alíeles may be present in this GD subpopulation.

Integrable plasmids pGR102 and pWR103 containing ribosomal sequences from within the transcriptional units for 16S and 23S were used to transform Bacillus subtilis. To date, these plasmids integrated into 7 of 10 known rrn operons. Two such events occurred at unassigned operons, revealing a close linkage of the CAT gene of the plasmid to pha-1 situated between dal-) and purB33 for rrnE and to thiA78 situated between glyB133 and tre-12 for rrnD. All seven integration events that led to the loss of unique ribosomal BclI fragments can now be assigned to known rrn operons.In the genome of Bacillus subtilis there are as many as 10 rRNA gene sets clustered in three groups (5). A major group is composed of rrnO and rrnA located at the replication origin (24,26,35) and the closely situated (5, 26) repeats (rrnI, rrnH, rrnG) found near the attachment site of phage SP02 (5). Minor clusters containing rrnB and rrnC have been mapped in the region between thr-S and aroG (5), and one cistron is believed to be located at the ilvBC-leu region (7,11,31). Strains of B. subtilis have been shown to contain only nine operons because of a deletion (11,18) or to exhibit rearrangements involving ribosomal sequences (3,11). Each transcriptional unit is arranged in the order 16S, 23S, and 5S rRNA genes, with two to three operons (including rrnO and rrnA) containing tRNAAla and tRNAlie genes in the abutment region between the 16S and 23S rDNA sequences (18, 33). We employed the integrative mapping method of Haldenwang et al. (13) to ascertain whether integrations can occur in operons, to locate various unmapped rrn genes on the chromosome, and to determine the physiological effects on growth and sporulation of such events in B. subtilis. In this study, the bifunctional plasmid pJH101 was used (10). The plasmid cannot replicate in B. subtilis because it does not contain an origin of replication for B. subtilis. If a region of homology with the B. subtilis chromosome is inserted in pJH101, the resultant plasmid gains the ability to transform competent cells and also, on occasion, integrates by Campbell-like recombination (10). We subcloned two similar but not identical ribosomal fragments from within the transcriptional unit for 16S and 23S which originated from the operons rrnO and rrnI. The resultant plasmids pGR102 and pWR103 can theoretically integrate into any of the 10 ribosomal operons by a Campbell-like insertion, leaving the entire plasmid with its antibiotic marker adjacent to the homologous region. We report here seven different integration events by these plasmids, two of which occur at unassigned operons (rrnD and rrnE), increasing the number from seven to nine known genomic locations in B. subtilis. containing integrated plasmids were constructed by competent cell transformation, verified for integration by hybridization, and checked for site of integration by PBS1 transduction mapping. We designated the various transformants as the B. subtilis strain Ql plasmids, with fl denoting plasmid integration. The plasmids with and wi...

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