Firing Up the Crucible with Senator Lee Metcalf & Governor Forrest Anderson -- Judge Gordon Bennett in “The Crucible of Change”

During the lead-up to Montana second progressive era, Lee Metcalf and Forrest Anderson, along with others, kept the progressive flame lit in Montana. Metcalf’s political history is replete with close electoral wins because of his commitment to progressive ideals when the times were not always politically favorable for that. As State Legislator, MT Supreme Court Justice, Congressman and eventually as US Senator, Lee won races by as little as 55 votes because he stuck to his guns as a progressive. In Forrest Anderson’s career as a County Attorney, State Legislator, MT Supreme Court Justice and 12 years as MT Attorney General he was respected as a pragmatic practitioner of politics. But during that entire career leading up to his election as Governor, Forrest Anderson was also a stalwart supporter of the progressive agenda exemplified by FDR and the New Deal, which brought folks out of the Great Depression that was brought on by the bad policies of the GOP and big business. As MT’s second progressive period began in 1965, the first important election was Senator Metcalf’s successful re-election battle in 1966 with the sitting MT Governor, Tim Babcock. And the progressive express was really ignited by the election of Forrest Anderson as Governor in 1968 after 16 years of Republican Governors in MT. Gordon Bennett played a rather unique role, being a confidant of Metcalf and Anderson, both who respected his wide and varied experience, his intellect, and his roots in progressivism beginning with his formative years in the Red Corner of NE Montana. Working with Senator Metcalf and his team, including Brit Englund, Vic Reinemer, Peggy McLaughlin, Betty Davis and Jack Condon among others, Bennett helped shape the progressive message both in Washington DC and MT. Progressive labor and farm organizations, part of the progressive coalition, benefitted from Bennett’s advice and counsel and aided the Senator in his career including the huge challenge of having a sitting popular governor run against him for the Senate in 1966. Metcalf’s noted intern program produced a cadre of progressive leaders in Montana over the years. Most notably, Ron Richards transitioned from Metcalf Intern to Executive Secretary of the Montana Democratic Party (MDP) and assisted, along with Bennett, in the 1966 Metcalf-Babcock race in a big way. As Executive Secretary Richards was critical to the success of the MDP as a platform for Forrest Anderson’s general election run and win in 1968. After Forrest’s gubernatorial election, Richards became Executive Assistant (now called Chief of Staff) for Governor Anderson and also for Governor Thomas Judge. The Metcalf progressive strain, exemplified by many including Richards and Bennett, permeated Democratic politics during the second progressive era. So, too, did the coalition that supported Metcalf and his policies. The progressivism of the period of “In the Crucible of Change” was fired up by Lee Metcalf, Forrest Anderson and their supporters and coalitions, and Gordon Bennett was in the center of all of that, helping fire up the crucible, setting the stage for many policy advancements in both Washington DC and Montana. Gordon Bennett’s important role in the 1966 re-election of Senator Lee Metcalf and the 1968 election of Governor Forrest Anderson, as well as his wide experience in government and politics of that time allows him to provide us with an insider’s personal perspective of those races and other events at the beginning of the period of progressive change being documented “In the Crucible of Change,” as well as his personal insights into the larger political/policy picture of Montana. Gordon Bennett, a major and formative player “In the Crucible of Change,” was born in the far northeast town of Scobey, MT in 1922. He attended school in Scobey through the eighth grade and graduated from Helena High School. After attending Carroll College for two years, he received his BA in economics from Carleton College in Northfield, MN. During a brief stint on the east coast, his daily reading of the New York Times (“best newspaper in the world at that time … and now”) inspired him to pursue a career in journalism. He received his MA in Journalism from the University of Missouri and entered the field. As a reporter for the Great Falls Tribune under the ownership and management of the Warden Family, he observed and competed with the rigid control of Montana’s press by the Anaconda Company (the Great Falls Tribune was the only large newspaper in Montana NOT owned by ACM). Following his intellectual curiosity and his philosophical bend, he attended a number of Farm-Labor Institutes which he credits with motivating him to pursue solutions to economic and social woes through the law. In 1956, at the age of 34, he received his Juris Doctorate degree from the Georgetown University Law Center in Washington, DC. Bennett’s varied career included eighteen years as a farmer, four years in the US Army during WWII (1942-46), two years as Assistant MT Attorney General (1957-59) with Forrest Anderson, three years in private practice in Glasgow (1959-61), two years as Associate Solicitor in the Department of Interior in Washington, DC (1961-62), and private law practice in Helena from 1962 to 1969. While in Helena he was an unsuccessful candidate for the Montana Supreme Court (1962) and cemented his previous relationships with Attorney General Forrest Anderson and US Senator Lee Metcalf. Bennett modestly refuses to accept the title of Campaign Manager for either Lee Metcalf (1966 re-election over the challenger, MT Republican Governor Tim Babcock) or Forrest Anderson (his 1968 election as Governor), saying that “they ran their campaigns … we were only there to help.” But he has been generally recognized as having filled that critical role in both of those critical elections. After Governor Anderson’s election in 1968, Bennett was appointed Director of the MT Unemployment Compensation Commission, a position from where he could be a close advisor and confidant of the new Governor. In 1971, Governor Anderson appointed him Judge in the most important jurisdiction in Montana, the 1st Judicial District in Helena, a position he held for seventeen years (1971-88). Upon stepping down from his judgeship, for twenty years (1988-2008) he was a law instructor, mediator and arbitrator. He currently resides in Helena with his wife, Norma Tirrell, former newspaper reporter and researcher/writer. Bennett has two adult children and four grandchildren.

Naturalistic study on the effectiveness of psycho-oncological interventions in cancer patients and their partners

BACKGROUND There is evidence for the efficacy of psycho-oncological interventions (POI) in randomized controlled trials for cancer patients. Our objective was to explore, under naturalistic conditions (using propensity score matching), whether POI are effective to decrease anxiety, depression, distress and overall psychopathological symptoms within cancer patients and their partners. METHODS This study was conducted in the Oncology and Hematology Center of a University clinic in Switzerland with a group of 186 patients and 117 partners. Outcome measures of mental health were the Hospital Anxiety and Depression Scale and the Symptom Checklist (SCL-9-K). Repeated-measures ANOVAs were used to analyze change over time and group effects between individuals with POI vs. without POI. RESULTS Highly distressed patients and their partners participating in POI reported better mental health over time. Among moderately distressed patients, a decrease over time emerged in depression and distress independent of POI. No effectiveness of POI could be demonstrated in moderately distressed patients and partners. CONCLUSION Most of the highly distressed patients receive additional POI and therefore conclusions about the efficacy of POI are difficult. For moderately distressed individuals, POI as implemented in Switzerland does not improve mental health in such patients and their partners, which may be caused by very time limited POI treatments. Studies with more intense POI treatments are needed.

Molecular phylogenetic analyses identify Alpine differentiation and dysploid chromosome number changes as major forces for the evolution of the European endemic Phyteuma (Campanulaceae)

Phyteuma is a chromosomally and ecologically diverse vascular plant genus and constitutes an excellent system for studying both the role of chromosomal change for species diversification and the evolution of high-mountain biota. This kind of research is, however, hampered by the lack of a sound phylogenetic framework exacerbated by the notoriously low predictive power of traditional taxonomy with respect to phylogenetic relationships in Campanulaceae. Based on a comprehensive taxon sampling and analyses of nuclear and plastid sequence and AFLP fingerprint data, Phyteuma is confirmed as a monophyletic group sister to the monotypic Physoplexis, which is in line with their peculiar flower morphologies. Within Phyteuma two clades, largely corresponding to previously recognized sections, are consistently found. The traditional circumscription of taxonomic series is largely rejected. Whereas distinctness of the currently recognized species is mostly corroborated, some interspecific relationships remain ambiguous due to incongruences between nuclear and plastid data. Major forces for diversification and evolution of Phyteuma are descending dysploidy (i.e., a decrease in chromosome base number) as well as allopatric and ecological differentiation within the Alps, the genus' center of species diversity. (C) 2013 Elsevier Inc. All rights reserved.

Information Integration Theory and the Construction of Individually Perceived Group Efficacy

According to Bandura (1997) efficacy beliefs are a primary determinant of motivation. Still, very little is known about the processes through which people integrate situational factors to form efficacy beliefs (Myers & Feltz, 2007). The aim of this study was to gain insight into the cognitive construction of subjective group-efficacy beliefs. Only with a sound understanding of those processes is there a sufficient base to derive psychological interventions aimed at group-efficacy beliefs. According to cognitive theories (e.g., Miller, Galanter, & Pribram, 1973) individual group-efficacy beliefs can be seen as the result of a comparison between the demands of a group task and the resources of the performing group. At the center of this comparison are internally represented structures of the group task and plans to perform it. The empirical plausibility of this notion was tested using functional measurement theory (Anderson, 1981). Twenty-three students (M = 23.30 years; SD = 3.39; 35 % females) of the University of Bern repeatedly judged the efficacy of groups in different group tasks. The groups consisted of the subjects and another one to two fictive group members. The latter were manipulated by their value (low, medium, high) in task-relevant abilities. Data obtained from multiple full factorial designs were structured with individuals as second level units and analyzed using mixed linear models. The task-relevant abilities of group members, specified as fixed factors, all had highly significant effects on subjects’ group-efficacy judgments. The effect sizes of the ability factors showed to be dependent on the respective abilities’ importance in a given task. In additive tasks (Steiner, 1972) group resources were integrated in a linear fashion whereas significant interaction between factors was obtained in interdependent tasks. The results also showed that people take into account other group members’ efficacy beliefs when forming their own group-efficacy beliefs. The results support the notion that personal group-efficacy beliefs are obtained by comparing the demands of a task with the performing groups’ resources. Psychological factors such as other team members’ efficacy beliefs are thereby being considered task relevant resources and affect subjective group-efficacy beliefs. This latter finding underlines the adequacy of multidimensional measures. While the validity of collective efficacy measures is usually estimated by how well they predict performances, the results of this study allow for a somewhat internal validity criterion. It is concluded that Information Integration Theory holds potential to further help understand people’s cognitive functioning in sport relevant situations.

Lateral incisor agenesis, canine impaction and characteristics of supernumerary teeth in a South European male population

Objective: To assess the prevalence of lateral incisor agenesis impacted canines and supernumerary teeth in a young adult male population. Materials and Methods: The panoramic radiographs of 1745 military students (mean age: 18.6 ± 0.52 years) who attended the Center of Aviation Medicine of the Armed Forces of Greece during the period 1997-2011 were initially analyzed for lateral incisor agenesis by two observers. After exclusion of the known orthodontic cases, a subgroup of 1636 examinees (mean age: 18.6 ± 0.44 years) was evaluated for canine impaction and supernumerary teeth. Results: Twenty-eight missing lateral incisors were observed in 22 military students, indicating an incidence of 1.3% in the investigated population. No lateral incisor agenesis was detected in the mandibular arch. A prevalence rate of 0.8% was determined for canine impaction in the sample of young adults. The majority of impacted teeth (86.7%) were diagnosed in the maxillary arch. Thirty-five supernumerary teeth were observed in 24 examinees (prevalence rate: 1.5%). The ratio of supernumerary teeth located in the maxilla versus the mandible was 2.2:1. The most common type of supernumerary tooth was the upper distomolar. Conclusion: The prevalence of lateral incisor agenesis, canine impaction, and supernumerary teeth ranged from 0.8 to 1.5% in the sample of male Greek military students.

Papain-induced in vitro disc degeneration model for the study of injectable nucleus pulposus therapy

BACKGROUND CONTEXT Proteolytic enzyme digestion of the intervertebral disc (IVD) offers a method to simulate a condition of disc degeneration for the study of cell-scaffold constructs in the degenerated disc. PURPOSE To characterize an in vitro disc degeneration model (DDM) of different severities of glycosaminoglycans (GAG) and water loss by using papain, and to determine the initial response of the human mesenchymal stem cells (MSCs) introduced into this DDM. STUDY DESIGN Disc degeneration model of a bovine disc explant with an end plate was induced by the injection of papain at various concentrations. Labeled MSCs were later introduced in this model. METHODS Phosphate-buffered saline (PBS control) or papain in various concentrations (3, 15, 30, 60, and 150 U/mL) were injected into the bovine caudal IVD explants. Ten days after the injection, GAG content of the discs was evaluated by dimethylmethylene blue assay and cell viability was determined by live/dead staining together with confocal microscopy. Overall matrix composition was evaluated by histology, and water content was visualized by magnetic resonance imaging. Compressive and torsional stiffness of the DDM were also recorded. In the second part, MSCs were labeled with a fluorescence cell membrane tracker and injected into the nucleus of the DDM or a PBS control. Mesenchymal stem cell viability and distribution were evaluated by confocal microscopy. RESULTS A large drop of GAG and water content of the bovine disc were obtained by injecting >30 U/mL papain. Magnetic resonance imaging showed Grade II, III, and IV disc degeneration by injecting 30, 60, and 150 U/mL papain. A cavity in the center of the disc could facilitate later injection of the nucleus pulposus tissue engineering construct while retaining an intact annulus fibrosus. The remaining disc cell viability was not affected. Mesenchymal stem cells injected into the protease-treated DDM disc showed significantly higher cell viability than when injected into the PBS-injected control disc. CONCLUSIONS By varying the concentration of papain for injection, an increasing amount of GAG and water loss could be induced to simulate the different severities of disc degeneration. MSC suspension introduced into the disc has a very low short-term survival. However, it should be clear that this bovine IVD DDM does not reflect a clinical situation but offers exciting possibilities to test novel tissue engineering protocols.

In situ dehydration behavior of zeolite-like pentagonite: A single-crystal X-ray study

The structural modifications upon heating of pentagonite, Ca(VO)(Si4O10)·4H2O (space group Ccm21, a=10.3708(2), b=14.0643(2), c=8.97810(10) Å, V=1309.53(3) Å3) were investigated by in situ temperature dependent single-crystal X-ray structure refinements. Diffraction data of a sample from Poona district (India) have been measured in steps of 25 up to 250 °C and in steps of 50 °C between 250 and 400 °C. Pentagonite has a porous framework structure made up by layers of silicate tetrahedra connected by V4+O5 square pyramids. Ca and H2O molecules are extraframework occupants. Room temperature diffraction data allowed refinement of H positions. The hydrogen-bond system links the extraframework occupants to the silicate layers and also interconnects the H2O molecules located inside the channels. Ca is seven-fold coordinated forming four bonds to O of the tetrahedral framework and three bonds to extraframework H2O. The H2O molecule at O9 showing a high displacement parameter is not bonded to Ca. The dehydration in pentagonite proceeds in three steps. At 100 °C the H2O molecule at O8 was released while O9 moved towards Ca. As a consequence the displacement parameter of H2O at O9 halved compared to that at room temperature. The unit-cell volume decreased to 1287.33(3) Å3 leading to a formula with 3H2O per formula unit (pfu). Ca remained seven-fold coordinated. At 175 °C Ca(VO)(Si4O10)·3H2O transformed into a new phase with 1H2O molecule pfu characterized by doubling of the c axis and the monoclinic space group Pn. Severe bending of specific TOT angles led to contraction of the porous three-dimensional framework. In addition, H2O at O9 was expelled while H2O at O7 approached a position in the center of the channel. The normalized volume decreased to 1069.44(9) Å3. The Ca coordination reduced from seven- to six-fold. At 225 °C a new anhydrous phase with space group Pna21 but without doubling of c had formed. Release of H2O at O7 caused additional contraction of TOT angles and volume reduction (V=1036.31(9) Å3). Ca adopted five-fold coordination. During heating excursion up to 400 °C this anhydrous phase remained preserved. Between room temperature and 225 °C the unit-cell volume decreased by 21% due to dehydration. The dehydration steps compare well with the thermo-gravimetric data reported in the literature.

Earth gravity field recovery using GPS, GLONASS, and SLR satellites

The time variable Earth’s gravity field provides the information about mass transport within the system Earth, i.e., the relationship of mass transport between atmosphere, oceans, and land hydrology. We recover the low-degree parameters of the time variable gravity field using microwave observations from GPS and GLONASS satellites and from SLR data to five geodetic satellites, namely LAGEOS-1/2, Starlette, Stella, and AJISAI. GPS satellites are particularly sensitive to specific coefficients of the Earth's gravity field, because of the deep 2:1 orbital resonance with Earth rotation (two revolutions of the GPS satellites per sidereal day). The resonant coefficients cause, among other, a “secular” drift (actually periodic variations of very long periods) of the semi-major axes of up to 5.3 m/day of GPS satellites. We processed 10 years of GPS and GLONASS data using the standard orbit models from the Center of Orbit Determination in Europe (CODE) with a simultaneous estimation of the Earth gravity field coefficients and other parameters, e.g., satellite orbit parameters, station coordinates, Earth rotation parameters, troposphere delays, etc. The weekly GNSS gravity solutions up to degree and order 4/4 are compared to the weekly SLR gravity field solutions. The SLR-derived geopotential coefficients are compared to monthly GRACE and CHAMP results.

Empirical chemosensitivity testing in a spheroid model of ovarian cancer using a microfluidics-based multiplex platform.

The use of biomarkers to infer drug response in patients is being actively pursued, yet significant challenges with this approach, including the complicated interconnection of pathways, have limited its application. Direct empirical testing of tumor sensitivity would arguably provide a more reliable predictive value, although it has garnered little attention largely due to the technical difficulties associated with this approach. We hypothesize that the application of recently developed microtechnologies, coupled to more complex 3-dimensional cell cultures, could provide a model to address some of these issues. As a proof of concept, we developed a microfluidic device where spheroids of the serous epithelial ovarian cancer cell line TOV112D are entrapped and assayed for their chemoresponse to carboplatin and paclitaxel, two therapeutic agents routinely used for the treatment of ovarian cancer. In order to index the chemoresponse, we analyzed the spatiotemporal evolution of the mortality fraction, as judged by vital dyes and confocal microscopy, within spheroids subjected to different drug concentrations and treatment durations inside the microfluidic device. To reflect microenvironment effects, we tested the effect of exogenous extracellular matrix and serum supplementation during spheroid formation on their chemotherapeutic response. Spheroids displayed augmented chemoresistance in comparison to monolayer culturing. This resistance was further increased by the simultaneous presence of both extracellular matrix and high serum concentration during spheroid formation. Following exposure to chemotherapeutics, cell death profiles were not uniform throughout the spheroid. The highest cell death fraction was found at the center of the spheroid and the lowest at the periphery. Collectively, the results demonstrate the validity of the approach, and provide the basis for further investigation of chemotherapeutic responses in ovarian cancer using microfluidics technology. In the future, such microdevices could provide the framework to assay drug sensitivity in a timeframe suitable for clinical decision making.

Characterization of Optically Stimulated Luminescent Detectors in Photon & Proton Beams for Use in Anthropomorphic Phantoms

This study investigated characteristics of optically stimulated luminescent detectors (OSLDs) in protons, allowing comparison to thermoluminescent detectors, and to be implemented into the Radiological Physics Center’s (RPC) remote audit quality assurance program for protons, and for remote anthropomorphic phantom irradiations. The OSLDs used were aluminum oxide (Al2O3:C) nanoDots from Landauer, Inc. (Glenwood, Ill.) measuring 10x10x2 mm3. A square, 20(L)x20(W)x0.5(H) cm3 piece of solid water was fabricated with pockets to allow OSLDs and TLDs to be irradiated simultaneously and perpendicular to the beam. Irradiations were performed at 5cm depth in photons, and in the center of a 10 cm SOBP in a 200MeV proton beam. Additionally, the Radiological Physics Center’s anthropomorphic pelvic phantom was used to test the angular dependence of OSLDs in photons and protons. A cylindrical insert in the phantom allows the dosimeters to be rotated to any angle with a fixed gantry angle. OSLDs were irradiated at 12 angles between 0 and 360 degrees. The OSLDs were read out with a MicroStar reader from Landauer, Inc. Dose response indicates that at angles where the dosimeter is near parallel with the radiation beam response is reduced slightly. Measurements in proton beams do not show significant angular dependence. Post-irradiation fading of OSLDs was studied in proton beams to determine if the fading was different than that of photons. The fading results showed no significant difference from results in photon beams. OSLDs and TLDs are comparable within 3% in photon beams and a correction factor can be posited for proton beams. With angular dependence characteristics defined, OSLDs can be implemented into multiple-field treatment plans in photons and protons and used in the RPC’s quality assurance program.

Impact of a stress coping strategy on perceived stress levels and performance during a simulated cardiopulmonary resuscitation: a randomized controlled trial

Background Cardiopulmonary resuscitation (CPR) causes significant stress, which may cause deficiencies in attention and increase distractibility. This may lead to misjudgements of priorities and delays in CPR performance, which may further increase mental stress (vicious cycle). This study assessed the impact of a task-focusing strategy on perceived stress levels and performance during a simulated CPR scenario. Methods This prospective, randomized-controlled trial was conducted at the simulator-center of the University Hospital Basel, Switzerland. A total of 124 volunteer medical students were randomized to receive a 10 minute instruction to cope with stress by loudly posing two taskfocusing questions (“what is the patient’s condition?”, “what immediate action is needed?”) when feeling overwhelmed by stress (intervention group) or a control group. The primary outcome was the perceived levels of stress and feeling overwhelmed (stress/overload); secondary outcomes were hands-on time, time to start CPR and number of leadership statements. Results Participants in the intervention group reported significantly less stress/overload levels compared to the control group (mean difference: -0.6 (95%CI −1.3, -0.1), p=0.04). Higher stress/overload was associated with less hands-on time. Leadership statements did not differ between groups, but the number of leadership statements did relate to performance. Hands-on time was longer in the intervention- group, but the difference was not statistically different (difference 5.5 (95%CI −3.1, 14.2), p=0.2); there were no differences in time to start CPR (difference −1.4 (95%CI −8.4, 5.7), p=0.71). Conclusions A brief stress-coping strategy moderately decreased perceived stress without significantly affecting performance in a simulated CPR. Further studies should investigate more intense interventions for reducing stress.

A biophysical model of synaptic plasticity and metaplasticity can account for the dynamics of the backward shift of hippocampal place fields.

Hippocampal place cells in the rat undergo experience-dependent changes when the rat runs stereotyped routes. One such change, the backward shift of the place field center of mass, has been linked by previous modeling efforts to spike-timing-dependent plasticity (STDP). However, these models did not account for the termination of the place field shift and they were based on an abstract implementation of STDP that ignores many of the features found in cortical plasticity. Here, instead of the abstract STDP model, we use a calcium-dependent plasticity (CaDP) learning rule that can account for many of the observed properties of cortical plasticity. We use the CaDP learning rule in combination with a model of metaplasticity to simulate place field dynamics. Without any major changes to the parameters of the original model, the present simulations account both for the initial rapid place field shift and for the subsequent slowing down of this shift. These results suggest that the CaDP model captures the essence of a general cortical mechanism of synaptic plasticity, which may underlie numerous forms of synaptic plasticity observed both in vivo and in vitro.

The effect of shoulder variation on IMRT and SmartArc for Head and Neck Cancer

Purpose: First, to determine an average and maximum displacement of the shoulder relative to isocenter over the course of treatment. Second, to establish the dosimetric effect of shoulder displacements relative to correct isocenter alignment on the dose delivered to the target and the surrounding structures for head and neck cancer patients. Method and Materials: The frequency of shoulder shifts of various magnitudes relative to isocenter was assessed for 4 patients using image registration software. The location of the center of the right and left humeral head relative to isocenter (usually C2) was found daily from CT on rails scans, and was compared to the location of the humeral heads relative to isocenter on the initial simulation CT. Three Baseline head and neck IMRT and SmartArc plans were generated in Pinnacle based on simulation CTs. The CT datasets (external contour and boney structures) were then modified to represent shifts of the shoulder (relative to isocenter) between 3 mm and 15 mm in the SI, AP, and LR directions. The initial plans were recalculated on the image sets with shifted shoulders. Results: On average, shoulder variation was 2-5 mm in each direction, although displacements of over 1 cm in the inferior and posterior directions occurred. Shoulder shifts induced perturbations in the dose distribution, although generally only for large shifts. Most substantially, large, superior shifts resulted in coverage loss by the 95% isodose line for targets in the lower neck. Inferior shifts elevated the dose to the brachial plexus by 0.6-4.1 Gy. SmartArc plans showed similar loss of target coverage as IMRT plans. Conclusions: The position of the shoulder can have an impact on target coverage and critical structure dose. Shoulder position may need to be considered for setup of head and neck patients depending on target location.

Functions and intramolecular interactions of ribosomal RNA in decoding of termination codons

Two regions in the 3$\prime$ domain of 16S rRNA (the RNA of the small ribosomal subunit) have been implicated in decoding of termination codons. Using segment-directed PCR random mutagenesis, I isolated 33 translational suppressor mutations in the 3$\prime$ domain of 16S rRNA. Characterization of the mutations by both genetic and biochemical methods indicated that some of the mutations are defective in UGA-specific peptide chain termination and that others may be defective in peptide chain termination at all termination codons. The studies of the mutations at an internal loop in the non-conserved region of helix 44 also indicated that this structure, in a non-conserved region of 16S rRNA, is involved in both peptide chain termination and assembly of 16S rRNA.^ With a suppressible trpA UAG nonsense mutation, a spontaneously arising translational suppressor mutation was isolated in the rrnB operon cloned into a pBR322-derived plasmid. The mutation caused suppression of UAG at two codon positions in trpA but did not suppress UAA or UGA mutations at the same trpA positions. The specificity of the rRNA suppressor mutation suggests that it may cause a defect in UAG-specific peptide chain termination. The mutation is a single nucleotide deletion (G2484$\Delta$) in helix 89 of 23S rRNA (the large RNA of the large ribosomal subunit). The result indicates a functional interaction between two regions of 23S rRNA. Furthermore, it provides suggestive in vivo evidence for the involvement of the peptidyl-transferase center of 23S rRNA in peptide chain termination. The $\Delta$2484 and A1093/$\Delta$2484 (double) mutations were also observed to alter the decoding specificity of the suppressor tRNA lysT(U70), which has a mutation in its acceptor stem. That result suggests that there is an interaction between the stem-loop region of helix 89 of 23S rRNA and the acceptor stem of tRNA during decoding and that the interaction is important for the decoding specificity of tRNA.^ Using gene manipulation procedures, I have constructed a new expression vector to express and purify the cellular protein factors required for a recently developed, realistic in vitro termination assay. The gene for each protein was cloned into the newly constructed vector in such a way that expression yielded a protein with an N-terminal affinity tag, for specific, rapid purification. The amino terminus was engineered so that, after purification, the unwanted N-terminal tag can be completely removed from the protein by thrombin cleavage, yielding a natural amino acid sequence for each protein. I have cloned the genes for EF-G and all three release factors into this new expression vector and the genes for all the other protein factors into a pCAL-n expression vector. These constructs will allow our laboratory group to quickly and inexpensively purify all the protein factors needed for the new in vitro termination assay. (Abstract shortened by UMI.) ^