Armed services vocational aptitude battery predictors of entry-level radiography students' success.

This study assessed the Armed Services Vocational Aptitude Battery (ASVAB) predictors of student success in the diagnostic imaging course at the Air Force's School of Health Care Sciences. Regression analyses provided two significant predictors (p < 0.05) of student success as identified by final grade. Those predictors were the mechanical and electronics scores on the ASVAB. Also, both scores were significantly different (p < 0.05) among those who passed, washed back (passed but required excess time), and disenrolled. Because a significant number of individuals take the ASVAB, entry-level radiography programs should consider using these predictors for admissions decisions.

Identification of amino acids stabilizing the tetramerization of the single stranded DNA binding protein from Escherichia coli.

Mutating the histidine at position 55 present at the subunit interface of the tetrameric E. coli single stranded DNA binding (SSB) protein to tyrosine or lysine leads to cells which are UV- and temperature-sensitive. The defects of both ssbH55Y (ssb-1) and ssbH55K can be overcome by increasing protein concentration, with the ssbH55K mutation producing a less stable, readily dissociating protein whose more severe replication and repair phenotypes were less easily ameliorated by protein amplification. In this study we selected and analyzed E. coli strains where the temperature sensitivity caused by the ssbH55K mutation was suppressed by spontaneous mutations that changed the glutamine at position 76 or 110 to leucine. Using guanidinium chloride denaturation monitored by sedimentation diffusion equilibrium experiments in the analytical ultracentrifuge, we demonstrate that the double mutant SSBH55KQ76L and SSBH55KQ110L proteins form more stable homotetramers as compared to the SSBH55K single mutant protein although they are less stable than wild-type SSB. Additionally, the single mutant proteins SSBQ76L and SSBQ110L form tetramers which are more resistant to guanidinium denaturation than wild-type SSB protein.

Air Force School of Health Care Science's quality of life study.

This was an empirically based assessment of non-prior service students' quality of life in the Air Force's School of Health Care Sciences. Analysis provided five results: (1) The overall quality of life at the school was good. (2) The variables accounting for student unhappiness were dormitory unsuitability and the students not being in their top-three career choices. (3) Structural changes were required at the dormitories. (4) The desire to succeed and how to achieve that success were the most important interests for students. (5) Loved ones and student independence were the greatest indicators of motivation. The findings resulted in three immediate corrections and two long-term recommendations to improve students' quality of life. The two long-term recommendations were to have an educational psychologist intervene when students are having significant learning problems, and to alter the selection process for recruiting. Both immediate corrections and long-term recommendations are useful for sister services.

Predictors of students' success in the Air Force School of Health Care Sciences.

This study assessed the predictors for students' success in the 882d Training Group's (Air Force School of Health Care Sciences) allied health courses at Sheppard Air Force Base, Texas. Correlation and regression analyses provided many predictors of students' success. From these predictors, the researchers ascertained demarcation points to provide direction for instructors and students. Using these parameters, proactive intervention by instructors, students, and the Student Learning Center will improve the success of allied health students. Further research conducted by the 882d Training Group in evaluating predictors of success will yield a seamless evaluation process. This process will establish an improved proactive educational system enhancing the career field selection process and the quality of students' education, as well as decrease costs.

Predictors of students' success in the Medical Service Apprentice course.

This study assessed the predictors of students' success in the 383d Training Squadron's Medical Service Apprentice course at Sheppard Air Force Base, Texas. Correlation and regression analyses provided seven predictors of students' success. These predictors were the Armed Services Vocational Aptitude Battery Mechanical score, block 1 grade, block 3 grade, block 3 student hours, block 4 grade, block 4 student hours, and National Registry Emergency Medical Technician Test 1 grade. From these predictors, researchers ascertained demarcation points to provide direction for instructors and students. Using this guidance, proactive intervention by instructors and the Student Learning Center will improve the success of Medical Service Apprentice course students. Further research conducted by the 882d Training Group to evaluate predictors of success will yield a seamless evaluation process. This process will establish an improved proactive educational system enhancing the career field selection process and the quality of student education.

Analysis of ssb mutations in vivo implicates SSB protein in two distinct pathways of SOS induction and in recombinational DNA repair.

Site-directed mutations in the Escherichia coli ssb gene were tested for the ability to complement a chromosomal ssb deletion for viability, and only the ssb W54-->G mutation failed to do so at the pSC101 copy level. Non-aromatic amino acid substitutions for SSB Trp-54 (ssb W54-->L and ssb W54-->S) produced the greatest effects on in vivo protein function including altered marker linkage subsequent to generalized transduction, extreme UV sensitivity, and a lack of ability to support SOS induction. Additionally, the ssb-113 (ssb P176-->S) mutation demonstrated the existence of both uvrA-dependent and uvrA-independent components of SOS induction. Although nucleotide excision repair appeared unaffected by alterations in the SSB protein, the mutational analysis suggests a direct role for SSB in recombinational repair.

Viability and preliminary in vivo characterization of site-directed mutants of Escherichia coli single-stranded DNA-binding protein.

Site-directed mutations involving selected amino acids of Escherichia coli single-stranded DNA-binding protein (SSB) were tested for their in vivo functionality when introduced into a chromosomal ssb deletion strain on a plasmid. All mutants complemented the ssb deletion for viability when present on a pSC101 derivative. The generation time with ssbW54S doubled in comparison to the ssb+ control, and both the ssbW54S- and ssbH55K-containing strains exhibited temperature sensitivity. ssbH55K, ssbW54S, ssbW88T, and ssbH55Y (ssb-1) strains displayed reduced survival to ultraviolet irradiation, while ssbW40T and ssbF60L strains were comparable to the ssb+ control strain. This study represents the first investigation of the in vivo properties of ssb mutations constructed for in vitro analysis of DNA binding by SSB.

The role of oriT in tra-dependent enhanced recombination between mini-F-lac-oriT and lambda plac5.

Recombination between F42lac and lambda plac5 is typically 20- to 50-fold more efficient than recombination between chromosomal lac and lambda plac5. This enhancement of recombination is recBCD-dependent and requires the expression of genes from the tra regulon of the F factor. Also required is oriT, the origin of F factor conjugational transfer, which must be located in-cis to the cellular copy of lac. In this study we show that enhanced recombination is not supported by an oriT point mutant that reduces oriT function in conjugation. We also present evidence that the activation of oriT for recombination enhancement involves the same strand-specific nick that is required for conjugal DNA transfer. Although it is thought that the role of oriT in recombination enhancement is related to the facilitated entry of RecBCD enzyme into the DNA duplex, we were unable to detect any double-strand breakage at oriT.

The single-stranded-DNA-binding protein encoded by the Escherichia coli F factor can complement a deletion of the chromosomal ssb gene.

Genes encoding single-stranded-DNA-binding proteins (SSBs) are carried by a variety of large self-transmissible plasmids, and it previously has been shown that these plasmid-borne genes can complement conditional lethal alleles of the ssb gene on the Escherichia coli chromosome for cellular viability. We have tested one of the plasmid-borne ssb genes, the ssf gene from the E. coli F factor, for its ability to complement total deletion of the chromosomal ssb gene for viability. We have found that ssf can complement the ssb deletion, but only when it is present on a high-copy-number plasmid. Cells that are totally dependent on the F-factor-encoded SSB for viability manifest growth properties indicative of problems in DNA replication.

traY and traI are required for oriT-dependent enhanced recombination between lac-containing plasmids and lambda plac5.

Recombination between F42lac and lambda plac5 is typically 20- to 50-fold more efficient than recombination between chromosomal lac and lambda plac5. This enhancement of recombination requires trans-acting factors located in the promoter-distal and promoter-proximal regions of the main traY-to-traI (traZ) operon. By testing the ability of deletion mutants of tra to support enhanced recombination, we have identified traY as the only product has been ruled out. We also report that traI is the only gene from the promoter-distal end of the traY to traI operon that is required for recombination enhancement. Of the two proposed domains of traI, we conclude that the oriT-nicking activity is essential, whereas the helicase activity is largely dispensable. The possibility of a third traI activity is also discussed.

Chromosomal transformation in the cyanobacterium Agmenellum quadruplicatum.

Chromosomal transformation of Agmenellum quadruplicatum PR-6 (= Synechococcus sp. strain 7002) was characterized for phenotypic expression, for exposure time to DNA, and for dependence on DNA concentration with regard to Rifr donor DNA. Exponentially growing cells of PR-6 were competent for chromosomal transformation. Competence decreased in cells in the stationary phase of growth or in cells deprived of a nitrogen source. Dark incubation of cells before exposure to donor DNA also decreased competence. Homologous Rifr and Strr DNA and heterologous Escherichia coli W3110 DNA were used in DNA-DNA competition studies, which clearly showed that DNA binding by PR-6 was nonspecific. DNA binding and uptake by PR-6 exhibited single-hit kinetics. Single-stranded DNA failed to transform competent cells of PR-6, and DNA eclipse was not observed, suggesting that double-stranded DNA was the substrate for the binding and uptake reactions during the transformation of PR-6. A significant improvement in transformation frequency was achieved by increasing the nitrate content of the culture medium and by lowering the temperature at which cells were exposed to donor DNA from 39 degrees C (the optimal temperature for growth) to 30 degrees C.

Nucleotide sequence and further characterization of the Synechococcus sp. strain PCC 7002 recA gene: complementation of a cyanobacterial recA mutation by the Escherichia coli recA gene.

The nucleotide sequence and transcript initiation site of the Synechococcus sp. strain PCC 7002 recA gene have been determined. The deduced amino acid sequence of the RecA protein of this cyanobacterium is 56% identical and 73% similar to the Escherichia coli RecA protein. Northern (RNA) blot analysis indicates that the Synechococcus strain PCC 7002 recA gene is transcribed as a monocistronic transcript 1,200 bases in length. The 5' endpoint of the recA mRNA was mapped by primer extension by using synthetic oligonucleotides of 17 and 27 nucleotides as primers. The nucleotide sequence 5' to the mapped endpoint contained sequence motifs bearing a striking resemblance to the heat shock (sigma 32-specific) promoters of E. coli but did not contain sequences similar to the E. coli SOS operator recognized by the LexA repressor. An insertion mutation introduced into the recA locus of Synechococcus strain PCC 7002 via homologous recombination resulted in the formation of diploids carrying both mutant and wild-type recA alleles. A variety of growth regimens and transformation procedures failed to produce a recA Synechococcus strain PCC 7002 mutant. However, introduction into these diploid cells of the E. coli recA gene in trans on a biphasic shuttle vector resulted in segregation of the cyanobacterial recA alleles, indicating that the E. coli recA gene was able to provide a function required for growth of recA Synechococcus strain PCC 7002 cells. This interpretation is supported by the observation that the E. coli recA gene is maintained in these cells when antibiotic selection for the shuttle vector is removed.

Mutations in the bacteriophage lambda pL/oL region that spontaneously occur in plasmid pRPZ126.

In a previous study (Chen and Porter, 1988), we isolated spontaneous mutations in a test plasmid that had occurred under non-selective conditions and assigned them to 1 of 6 different categories or groups. The test plasmid, pRPZ126, is a pBR322 derivative containing the bacteriophage lambda immunity region with the cI857 allele so that plasmid-containing cells shifted to 42 degrees C survive only if the expression of the lambda kil gene is prevented by mutation. 75% of the total spontaneous mutations obtained fall into two of these groups where there is no readily detectable change in plasmid size. The two groups differ in that the plasmids from the group 4 mutations are missing a specific HincII site while the plasmids from the group 5 mutations had no detectable plasmid change whatsoever. In this study, we randomly selected ten group 4 mutants and ten group 5 mutants and sequenced the lambda pL/oL region of the mutant plasmid. Of the ten group 4 mutants (HincII site missing), five involved a 24- or 44-basepair deletion in the pL/oL region of the plasmid. The other five group 4 mutants and four of the ten group 5 mutants were A-T to G-C transitions in the pL/oL region. The remaining six group 5 mutants did not demonstrate any sequence change in the pL/oL region of the plasmids. 8 out of 9 of these transition mutations occurred next to the 3' end of 3 different 5'-PyGGNPuNTG-3' sequences in the lambda operator region, and this same sequence is found adjacent to the A-T to G-C transition hotspot in the lac operator region (Schaaper et al., 1986). The 9th mutation, where the A-T to G-C transition occurred one basepair away from the lambda operator, was adjacent to a very similar sequence.

Use of the Escherichia coli SSB gene to prevent bioreactor takeover by plasmidless cells.

Reactor takeover by plasmidless cells is a major problem encountered when producing proteins from plasmid-borne genes in genetically engineered bacteria. We have approached this problem by deleting the essential ssb gene from the Escherichia coli chromosome and placing it on a plasmid. Plasmidless cells do not accumulate even after growing such strains under non-selective continuous culture conditions for extended periods of time. Other ssb-containing plasmids can be readily introduced into this E. coli strain by a plasmid-displacement technique. Using this system, we have achieved very high levels of beta-lactamase production in continuous culture without selective pressure.

Characterization of the types of mutational events that spontaneously occur in a plasmid system.

The immunity region from a cI857 derivative of bacteriophage lambda has been cloned into the EcoRI site of pBR322 to produce a plasmid that can be used to analyze spontaneous mutagenesis. Cells containing this plasmid are temperature-sensitive for growth unless mutations have occurred that somehow prevent the expression of the kil gene in the lambda fragment at non-permissive temperature. 678 such temperature-resistant mutants from 10 independent subcultures each of 2 different recA- E. coli strains have been collected, and the nature of the plasmid mutations obtained has been analyzed. All of the subcultures contained mutants that allowed growth at the restrictive temperature without showing a detectable change in plasmid size. 75% of the total mutants fell in this class. More than half of these mutations involved the lambda leftward promoter, pL, and such mutants were found in all 20 subcultures. The remaining 25% of the mutations involved a change in plasmid size and mutations of this class were found in 18 of 20 subcultures. 12% of the total mutants (found in 16 of 20 subcultures) had an insertion of IS1 in the region between pL and the lambda kil gene. 6% of the total mutants had undergone an IS1-mediated deletion, while 1% were mixed colonies in which multiple IS1-mediated events had occurred. About 1% of the total mutants had undergone complex IS1-mediated DNA rearrangement(s) that have not yet been characterized. In total, 11 of 20 subcultures yielded isolates where IS1-mediated rearrangements had occurred. The remaining 4% of the mutations included insertions of IS5, IS30, and an IS1 family member that appears to be IS1T as well as IS1T-mediated deletions and deletions that do not appear to have been mediated by any insertion sequence. A mutant with both an IS1 insertion and an alteration involving pL has also been isolated.

Mutations in the recD gene of Escherichia coli that raise the copy number of certain plasmids.

Chromosomal mutants were isolated in which, for several small plasmids, there was an increased amount of either covalently closed circular plasmid DNA or total plasmid DNA or both. The mutations were mapped to recD, which has been shown to affect exonuclease V activity and a variety of plasmid maintenance and replication functions. Our results suggest that rolling-circle plasmid replication can occur in recD mutants and that site-specific recombination can resolve the resulting linear multimers into covalently closed circular plasmid forms.

Molecular cloning and characterization of the recA gene from the cyanobacterium Synechococcus sp. strain PCC 7002.

The recA gene of Synechococcus sp. strain PCC 7002 was detected and cloned from a lambda gtwes genomic library by heterologous hybridization by using a gene-internal fragment of the Escherichia coli recA gene as the probe. The gene encodes a 38-kilodalton polypeptide which is antigenically related to the RecA protein of E. coli. The nucleotide sequence of a portion of the gene was determined. The translation of this region was 55% homologous to the E. coli protein; allowances for conservative amino acid replacements yield a homology value of about 74%. The cyanobacterial recA gene product was proficient in restoring homologous recombination and partial resistance to UV irradiation to recA mutants of E. coli. Heterologous hybridization experiments, in which the Synechococcus sp. strain PCC 7002 recA gene was used as the probe, indicate that a homologous gene is probably present in all cyanobacterial strains.

Heterospecific transformation among cyanobacteria.

Heterospecific transformation occurred between cyanobacteria currently classified in either the genus Synechococcus or Synechocystis. Cyanobacterial strains 73109 and 6906 were capable of physiological transformation.