Linkage analysis of the murine mos proto-oncogene on chromosome 4

A linkage analysis of the murine Mos gene, which codes for the c-mos proto-oncogene, was performed in 88 backcross progeny of an interspecies cross of laboratory mice and Mus spretus. Linkage was tested for four different genes on mouse chromosome 4: Aco-1, Mup-1, b, and Ifb. The gene order (from centromere) with intervening percentage recombination is Mos-15.9 (+/- 3.9)-Aco-1-5.6 (+/- 2.4)-Mup-1-3.4 (+/- 1.9)-b-5.6 (+/- 2.4)-Ifb. These results confirm the previous assignment of Mos to chromosome 4 on the basis of segregation in somatic cell hybrids (D. Swan et al., 1982, J. Virol. 44: 752-754) and show furthermore that Mos and the Ifa/Ifb clusters are not tightly linked as a group of intronless genes, but are separated by a map distance of 30.6 +/- 4.9 recombination units. The linkage data obtained in the present study place Mos in a region compatible with the physical map (D. W. Threadgill and J. E. Womack, 1988, Genomics 3: 82-86).     

The fast milk acidifying phenotype of Streptococcus thermophilus can be acquired by natural transformation of the genomic island encoding the cell-envelope proteinase PrtS

Background

FAD dependent glucose dehydrogenase (GDH) currently raises enormous interest in the field of glucose biosensors. Due to its superior properties such as high turnover rate, substrate specificity and oxygen independence, GDH makes its way into glucose biosensing. The recently discovered GDH from the ascomycete Glomerella cingulata is a novel candidate for such an electrochemical application, but also of interest to study the plant-pathogen interaction of a family of wide-spread, crop destroying fungi. Heterologous expression is a necessity to facilitate the production of GDH for biotechnological applications and to study its physiological role in the outbreak of anthracnose caused by Glomerella (anamorph Colletotrichum) spp.

Results

Heterologous expression of active G. cingulata GDH has been achieved in both Escherichia coli and Pichia pastoris, however, the expressed volumetric activity was about 4800-fold higher in P. pastoris. Expression in E. coli resulted mainly in the formation of inclusion bodies and only after co-expression with molecular chaperones enzymatic activity was detected. The fed-batch cultivation of a P. pastoris transformant resulted in an expression of 48,000 U L^-1 of GDH activity (57 mg L^-1). Recombinant GDH was purified by a two-step purification procedure with a yield of 71%. Comparative characterization of molecular and catalytic properties shows identical features for the GDH expressed in P. pastoris and the wild-type enzyme from its natural fungal source.

Conclusions

The heterologous expression of active GDH was greatly favoured in the eukaryotic host. The efficient expression in P. pastoris facilitates the production of genetically engineered GDH variants for electrochemical-, physiological- and structural studies.     

Linkage analysis of the murine interferon alpha locus (Ifa) on chromosome 4

We performed linkage analysis of the murine interferon alpha gene cluster, Ifa, with three different marker genes on chromosome 4: the major urinary protein locus (Mup-1), the brown locus (b) and the misty locus (m). The gene order (from centromere) with intervening percent recombination derived from a first three-point cross is Mup-1--13.6 (+/- 3.6)--Ifa--7.9 (+/- 2.8)--m and from a second three-point cross Mup-1--1.7 (+/- 1.7)--b--3.5 (+/- 2.4)--Ifa. The combined results indicate that the gene order, from the centromere, is Mup-1--b--Ifa--m.     

Development of a versatile procedure based on natural transformation for marker-free targeted genetic modification in Streptococcus thermophilus

A versatile natural transformation protocol was established for and successfully applied to 18 of the 19 Streptococcus thermophilus strains tested. The efficiency of the protocol enables the use of in vitro-amplified mutagenesis fragments to perform deletion or insertion of large genetic fragments. Depending on the phenotype linked to the mutation, markerless mutants can be selected either in two steps, i.e., resistance marker insertion and excision using an adapted Cre-loxP system, or in one step using a powerful positive screening procedure as illustrated here for histidine prototrophy.     

Hepatitis B Prevention in Small Rural Hospitals

A retrospective study of 246 potential hepatitis B exposure incidents in 12 rural hospitals in Arizona over a two-year period revealed a rate of 6.3 incidents per 100 employees per year. Needle punctures accounted for 68% of the incidents; 17% were cuts from instruments or broken glassware. Although 51% occurred in nursing personnel, housekeepers accounted for a surprising 19.5% of the reports. Only 50% of the employees received any medical attention following incidents. None received hepatitis B immune globulin (HBIG); seven received immune globulin (IG). The mean cost to the hospitals for the 122 incidents where treatment was given was $64.50 per incident. In all, 10 hospitals had no written policy for hepatitis B prevention, 3 did not stock IG and 11 did not stock HBIG. There was little awareness of hepatitis B as a nosocomial problem within these institutions, perhaps because no reported cases of clinical hepatitis B occurred in employees of the 12 hospitals in the two years.     

The Diarrhea of Travelers—Incidence in Foreign Students in the United States

To ascertain whether foreign visitors to the United States experience the diarrhea of travelers, 215 foreign students matriculating at the University of California, Los Angeles, were interviewed. The attack rate of diarrhea was 14.0 per cent during the first month after arrival in the United States. In a comparison group of 238 U.S. students, the attack rate of diarrhea in one month was 8.4 per cent. The difference in attack rate was not statistically significant.The diarrheal episodes reported by the foreign students were less abrupt in onset, less severe and of longer duration than the usual diarrhea of travelers. The typical severe explosive diarrhea of short duration usually described as “diarrhea of travelers” was not encountered among the foreign students in this study.The occurrence of diarrhea was not associated with the age, sex or race of the students, with the location of eating places, with the geographic area from which the foreign students came or with any characteristics of the trip to the United States. In the group of U.S. students there was an increased incidence of diarrhea among students who came from outside California to Los Angeles.