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Fructose-1,6-bisphosphatase (FBPase) deficiency is an autosomal recessive inherited disorder and may cause sudden unexpected infant death. We reported the first case of molecular diagnosis of FBPase deficiency, using cultured monocytes as a source for FBPase mRNA. In the present study, we confirmed the presence of the same genetic mutation in this patient by amplifying genomic DNA. Molecular analysis was also performed to diagnose another 12 Japanese patients with FBPase deficiency. Four mutations responsible for FBPase deficiency were identified in 10 patients from 8 unrelated families among a total of 13 patients from 11 unrelated families; no mutation was found in the remaining 3 patients from 3 unrelated families. The identified mutations included the mutation reported earlier, with an insertion of one G residue at base 961 in exon 7 (960/961insG) (10 alleles, including 2 alleles in the Japanese family from our previous report [46% of the 22 mutant alleles]), and three novel mutations--a G-->A transition at base 490 in exon 4 (G164S) (3 alleles [14%]), a C-->A transversion at base 530 in exon 4 (A177D) (1 allele [4%]), and a G-->T transversion at base 88 in exon 1 (E30X) (2 alleles [9%]). FBPase proteins with G164S or A177D mutations were enzymatically inactive when purified from E. coli. Another new mutation, a T-->C transition at base 974 in exon 7 (V325A), was found in the same allele with the G164S mutation in one family (one allele) but was not responsible for FBPase deficiency. Our results indicate that the insertion of one G residue at base 961 was associated with a preferential disease-causing alternation in 13 Japanese patients. Our results also indicate accurate carrier detection in eight families (73%) of 11 Japanese patients with FBPase deficiency, in whom mutations in both alleles were identified.

A method for effective elimination of drug resistance (R) and sex (F) factors in Escherichia coli K-12 strains by treatment with sodium dodecyl sulfate (SDS) is presented. Growth of E. coli harboring R or F factors in Penassay Broth containing SDS led to the loss of all or part of these genetic elements. Appearance of drug-susceptible or F-cells among survivors was observed after the culture reached the stationary phase. Drug-susceptible cells which had lost all of their resistance markers by SDS treatment could be efficiently infected with R or F factors. Among isolated segregants which came from resistant cells, tetracycline-susceptible cells were the major segregant class. Drug-susceptible cells gave no revertants to drug resistance. By treatment of F+ cells with SDS, unusual F+ cells which retained mating ability but showed resistance to M12 phage were also isolated, together with mutants of another type which lost mating ability but retained sensitivity to M12 phage. Since SDS is more toxic to R+ cells than R-cells, the isolation of drug-susceptible or F-cells under these conditions may be partly attributable to selective growth of drug-susceptible or F-cells in SDS-Penassay Broth.

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