• GABAergic neurons in the medullary raphé possess network independent chemosensitivity in situ

      Iceman, Kimberly E. (2012)
      The identity and location of central pH/CO2 sensitive chemoreceptors are not fully understood. Serotonin (5-HT) and γ-aminobutyric acid (GABA) synthesizing neurons in the medullary raphé have demonstrated intrinsic chemosensitivity in vitro. This evidence forms the basis for our "push-pull" model of raphé contributions to chemosensitivity. We have previously shown that CO2-stimulated 5-HT neurons occur in the medullary raphé in situ. Here, we test the hypothesis that the medullary raphé contains GABA synthesizing CO2-inhibited neurons that retain their chemosensitivity after pharmacological blockade of major fast synaptic inputs. To assess chemosensitivity, we record extracellular single neuron discharge during normocapnic and hypercapnic conditions within the medullary raphé of the unanesthetized juvenile rat in situ perfused decerebrate brainstem preparation. Network dependence of chemosensitivity is assessed by application of antagonists for AMPA, NMDA, glycine, and GABAa receptors that disrupt fast-synaptic network properties. Juxtacellular labeling and immunohistochemistry establish neurotransmitter phenotypes of recorded neurons. Results support independence of CO2-inhibited GABA neuron chemosensitivity from fast synaptic inputs.
    • Gardening With Annual Flowers In Interior Alaska

      Wagner, Pat; Griffith, Marilyn; Matheke, Grant (Agricultural and Forestry Experiment Station, School of Agriculture and Land Resources Management, University of Alaska-Fairbanks, 1987-04)
      This list of recommended annual flower varieties includes information on several hundred annual flower cultivars. The recommended varieties were selected from flowers grown in 1985 and 1986 at the Agricultural and Forestry Experiment Station Farm at the University of Alaska- Fairbanks. While this is by no means a complete listing of varieties suitable for Interior gardens, it does reflect many years of experience in annual flower production at the AFES farm. The methods used to evaluate the flowers and definitions for terms used in the listing can be found under Data Collection.
    • Gas and particle data from Fairbanks during the winter of 2019/2020

      Simpson, William; Cesler-Maloney, Meeta (N/A, 2020-09-30)
      This repository contains data from the relocatable air monitoring system (RAMS) trailer, which measured air pollution around sites in the Fairbanks North Star Borough (FNSB), Alaska. Measured pollutants were: SO2, CO, CO2, and particulate matter (PM). We additionally measured meteorological parameters such as temperatures, winds, and vertical differences in temperatures across an 11-meter tall tower.
    • Gaseous emissions from herding agent-mediated in-situ burning for Arctic oil spills

      Sartz, Patrik Pettersson; Aggarwal, Srijan; Barnes, David; Schnabel, William (2017-05)
      If a crude oil spill were to occur in partially ice-covered waters, many of the response tactics typically utilized in either open water or completely ice-covered conditions would become inefficient. In such situations, in-situ burning (ISB) can prove to be an efficient response tool; herding agent application is one available approach to thicken an oil slick. This study assessed the impacts on air quality following ISB tests on crude oil, in combination with herding agents, in partially ice-infested waters. The research focused on measuring downwind concentrations of respirable particulate matter (PM₂.₅) and seven different combustion gasses (CO, CO₂, NO, NO₂, NOx, SO₂, and VOCs) during five ISB events, with sampling instruments placed in-plume and 6-12 m away from the source area. The study also investigated if the utilized herding agent was detectable in the airborne plume. Findings include: 1) Concentrations of particulate matter (<2.5μm in diameter), SO₂, and CO were found to significantly (P <0.01) exceed various exposure limits and air quality standards, while the remaining compounds measured were significantly (P <0.01) below established exposure limits. Also, downwind, in the smoke plume, measured concentrations of SO₂, NOx, and total VOCs were higher than found in previous studies. It should be noted that instrument and methods not specifically approved by the U.S. Environmental Protection Agency, Occupational Safety and Health Administration, and the National Institute for Occupational Safety and Health were utilized during this study; 2) GC/MS analysis of aerosol samples collected utilizing a flow meter and carbon sorbent tubes in the smoke plume; the Siltech OP-40 silicone based functional group of the applied herding agent was not detected in the collected samples analyzed using GC/MS. Future research should include additional scalability studies where the concentrations of particulate matter and various combustion gasses are compared to modeled concentrations using computer software. Additional research is also needed to find a cost-effective method to decrease the amount of particulate matter during an in-situ burn. It is also recommended that guidance specific for conducting in-situ burns of crude oil or refined petroleum products in the Arctic is written and published by regulatory agencies, so the industry can rapidly make plans and propose such tactics if an incident did occur where mechanical or other non-mechanical response tactics are not feasible.
    • Gasser Wheat A New Grain for Alaska

      Taylor, R.L.; Brinsmade, J.C. (University of Alaska, Alaska Agricultural Experiment Station, 1955-10)
      Developed from the hybrid Diamond x Khogot by the Alaska Agricultural Experiment Station, Gasser wheat was released to seed producers in 1955. This variety exhibits an excellent combination of high yield and agronomic desirability m Alaska. It is superior to the standard variety Khogot in lodging resistance, shattering resistance, and grain quality. Gasser wheat is not satisfactory for commercial milling and baking. Its chief value in Alaska will be as a feed for poultry and livestock. Gasser is recommended for production in the Matanuska and Tanana Valleys when a wheat crop is desired. Later maturing than other cereals, Gasser must be planted early to insure maturity during the short growing season. Cultural recommendations are the same as for other cereals.
    • Gaussian process convolutions for Bayesian spatial classification

      Best, John K.; Short, Margaret; Goddard, Scott; Barry, Ron; McIntyre, Julie (2016-05)
      We compare three models for their ability to perform binary spatial classification. A geospatial data set consisting of observations that are either permafrost or not is used for this comparison. All three use an underlying Gaussian process. The first model considers this process to represent the log-odds of a positive classification (i.e. as permafrost). The second model uses a cutoff. Any locations where the process is positive are classified positively, while those that are negative are classified negatively. A probability of misclassification then gives the likelihood. The third model depends on two separate processes. The first represents a positive classification, while the second a negative classification. Of these two, the process with greater value at a location provides the classification. A probability of misclassification is also used to formulate the likelihood for this model. In all three cases, realizations of the underlying Gaussian processes were generated using a process convolution. A grid of knots (whose values were sampled using Markov Chain Monte Carlo) were convolved using an anisotropic Gaussian kernel. All three models provided adequate classifications, but the single and two-process models showed much tighter bounds on the border between the two states.
    • Gender Of Perpetrator, Gender Of Victim, And Relationship Between Perpetrator And Victim As Factors Influencing How Adults View Coercive Sexual Behavior In Childhood

      Bosek, Rebecca Lynn; Connor, Bill; Risley, Todd (2002)
      The sexual abuse of children by adults is a serious social problem. Some sexually abused children become sexually abusive toward others. This is sometimes called coercive sexual behavior, and little is known about how adults view these acts. A better understanding of how adults view coercive sexual behavior between children is critical due to the harm it causes victims, perpetrators, and society. Also, parents are typically held legally responsible for their minor children, and it is their responsibility to intervene in this type of behavior. Three hundred and eighty-five college students participated in a study that examined descriptions of coercive sexual behavior between elementary school-aged children. This study used a 2 x 2 x 2 factorial design to examine how gender of a child perpetrator, gender of a child victim, and relationship between a child perpetrator and child victim (peer or sibling) influence how adults view coercive sexual behavior in childhood. Participants read one of eight vignettes describing an incident of coercive sexual behavior between two children and answered a twenty-eight-item questionnaire based on it. Data was analyzed using correlation coefficients, factor analysis, and multivariate analysis of variance (MANOVA). Findings from the present study suggest that the gender of the children and the relationship between them are factors influencing how adults view coercive sexual behavior in childhood.
    • Gene by environment interactions between three candidate genes for obesity and omega-3 polyunsaturated fatty acids

      Pasker, Renee Leigh; Boyer, Bert; Wolf, Diane E.; Tiwari, Hemant (2008-12)
      Multi-factorial diseases, like obesity, are caused by genetic and environmental factors. Few studies examine potential interactions between genetic and environmental factors. Understanding these interactions can lead to better disease prevention. One important environmental factor related to obesity is omega-3 fatty acids. Yup’ik Eskimos consume a high amount of two omega-3 fatty acids, eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA). This study examined 10 single nucleotide polymorphisms (SNPs) in three genes: ADIPOQ, PPARG, and PPARGC1A. Also, 7 obesity phenotypes were examined: BMI, percent body fat, waist circumference, sum of skin-folds, plasma adiponectin, plasma triglycerides, and HDL cholesterol. Associations between these SNPs and phenotypes in 981 related Yup’ik Eskimos were examined using mixed models. Interactions were investigated with these SNPs and δ15N, a biomarker used to determine intake of EPA and DHA. The results showed that EPA and DHA modify the expression of all three genes. Additionally, SNPs in all three genes were associated with one or more obesity phenotypes. The most statistically significant results were with two SNPs in ADIPOQ and plasma adiponectin. This study supports the role of these genes in the etiology of obesity. Finally, this study demonstrates that these genes are influenced by EPA and DHA.
    • Gene-By-Diet Interactions And Obesity Among Yup'ik People Living In Southwest Alaska

      Lemas, Dominick; Boyer, Bert B.; O'Brien, Diane M.; Schulte, Marvin K.; Tiwari, Hemant K. (2012)
      BACKGROUND: Molecular approaches have expedited the discovery of human obesity genes, however the heritability explained by these loci remains low (<2%). Gene-by-environment interactions may partially account for the "missing heritability" attributed to variation in obesity phenotypes. OBJECTIVE: The specific aims of this dissertation were to (i) identify genetic polymorphisms associated with obesity-related phenotypes in Yup'ik people and (ii) evaluate how n-3 polyunsaturated fatty acid (n-3 PUFA) intake modifies associations between genetic polymorphisms and obesity-related phenotypes in a population with widely varying intake of n-3 PUFAs. APPROACH: We genotyped genetic polymorphisms in (1) candidate genes with a strong physiological role in obesity pathophysiology; (2) candidate genes identified in obesity whole-genome linkage studies that were regulated by n-3 PUFAs; and (3) candidate genes reproducibly implicated in obesity genome-wide association studies (GWAS). DATA & ANALYSES: We used Center for Alaska Native Health Research (CANHR) data collected between 2001 and 2008. We estimated dietary intake of n-3 PUFA using nitrogen stable isotope ratios (delta15N) of red blood cells (RBC) and obesity-related phenotypes were obtained by trained staff. Genotype-phenotype analyses used generalized linear models that accounted for familial correlations. RESULTS: Our analyses of candidate genes based on physiology revealed a polymorphism called P479L in carnitine palmitoyltransferase 1A (CPT1A) that was associated with elevated fasting HDL-cholesterol and all obesity phenotypes. Our investigation of candidate genes that are regulated by n-3 PUFAs and implicated in obesity whole-genome linkage studies demonstrate that polymorphisms in stearoyl CoA desaturase (SCD) and steroyl regulatory element binding protein (SLC2A4) were associated with obesity-related phenotypes; however n-3 PUFA intake did not modify associations between SCD and SLC2A4 polymorphisms and obesity phenotypes. Finally, our investigation of candidate genes reproducibly implicated in obesity GWAS demonstrated that genetic predisposition to obesity is associated with adiposity and that interactions with n-3 PUFA intake accounted for more than twice the phenotypic variation in adiposity. CONCLUSION: Taken together, results from this dissertation suggest that selecting candidate genes based on large-scale genomic analyses, such as linkage analyses and GWAS, has the potential to identify gene-by-environment interactions that partially account for the "missing heritability" attributed to obesity.
    • Genetic Ancestry Modeling And Performance Association In The Alaskan Sled Dog

      Huson, Heather Jay; Runstadler, Jonathan; Hundertmark, Kris; Ostrander, Elaine; Bailey-Wilson, Joan (2011)
      Alaskan sled dogs present us with the unique opportunity to study the development of a population of dogs produced from the selective breeding of high performance athletes. I establish that sled dogs are a genetically distinct population of dogs that segregate into two sub-groups based on their racing style of "sprint" or short distance and "distance" or long distance. The practice of interbreeding Alaskan sled dogs with various purebred dogs over the past century has allowed us to investigate the impact of these domestic breeds on the sled dog genome and their potential contribution to athletic performance. Here, I establish genetic profiles of both the sprint and the distance racing dogs using microsatellite-based markers, single-nucleotide polymorphism (SNP) arrays, and ancestry modeling. Population structure is assessed using clustering and principle component analyses. Inbreeding patterns are examined through population structure, inbreeding statistics, estimations of linkage disequilibrium, and autozygosity. Purebred breed components and their potential role in influencing performance attributes of Alaskan sled dogs were determined through genetic breed identification. Ancestry modeling was used to localize genomic regions of specific breed selection. These breeds were then analyzed for their genetic contribution to regions experiencing selection within the sprint or distance racing dogs. I determined regions of selective sweep and genome-wide association to the sprint or distance racing dogs. A genome-wide association analysis of heat tolerance performance in sprint dogs identified SNPs potentially regulating the MYH9 gene. This was the first genetic assessment of ancestry, inbreeding, and performance genes attributed to racing Alaskan sled dogs.
    • Genetic and chemical studies relating to α-aminoisobutyri acid (AIB) metabolism

      Bickmeier, Jeffrey A. (2004-12)
      The non-protein amino acid a-aminoisobutyric acid (Aib) occurs naturally in soil. Burkholderia cepacia utilizes Aib as a sole nitrogen source by virtue of a 2,2-dialkylglycine decarboxylase, which is encoded by the dgdA gene. In this study, 30 prokaryotes were cultured on minimal media containing Aib as the sole nitrogen source and identified by 16S rRNA gene sequencing. Several attempts were made to clone dgd genes from these organisms. Plasmid libraries were constructed in E. coli from DNA isolated from four Aib-utilizing bacteria. Libraries were screened for clones which demonstrated the ability to grow on Aib containing media and therefore contained cloned dgd genes. Sequence analysis of plasmids isolated from Aib utilizing clones revealed no similarity to dgd genes from B. cepacia. Also, 4-methyl-2-(4-nitrophenyl)-5(4H)-oxazolone was alkylated at the 4-position with various alkyl halides. This was the key step in synthesis of disubstituted N-p-nitrobenzoyl-DL-glycines, which are precursors to 2,2-dialkylglycines. Synthesis was performed in multiple steps. N-p-nitrobenzoyl-DL-alanine was synthesized by aminolysis of DL-alanine with benzoyl chloride. The 4-methyl-2-(4-nitrophenyl)-5(4H)-oxazolone was synthesized by dehydration of N-p-nitrobenzoyl-DL-alanine with acetic anhydride. Alkylation of 4-methyl-2-(4-nitrophenyl)-5(4H)-oxazolone was performed by adding base, which made the oxazolone enolate, and alkyl halide, which participated in SN2 reactions with the enolate. Substituted oxazolones were hydrolyzed into disubstituted N-p-nitrobenzoyl-DL-glycines with DCl.
    • Genetic And Environmental Effects On Developmental Timing, Otolith Formation, And Gill Raker Development In Pink Salmon From Auke Creek, Alaska

      Oxman, Dion; Gharrett, Anthony; Milo, Adkison,; Cailliet, Gregor; Hagen, Peter; Smoker, William (2012)
      To determine how inheritance, environment, and hybridization influenced developmental timing, otolith formation, and gill raker development in pink salmon (Oncorhynchus gorbuscha), full and half-sibling families from Auke Creek, Alaska and third generation outbred hybrids between Auke Creek females and Pillar Creek males from Kodiak Island, Alaska (1,000 km distant) were incubated in ambient, chilled, and warmed water. Variation in development time of embryos from the odd-year broodline was primarily influenced by additive genetic factors, whereas no genetic effect was detected in the even-year run. No genotype-by environment (GxE) effects were associated with sires or families in either broodline, indicating that the observed variation in development time was likely the result of phenotypic plasticity. Hybridization (outbreeding) significantly prolonged development time in both broodlines, indicating that the phenotypic effects of outbreeding can last at least three generations. Early otolith development was genetically conserved and canalized, but the phenotypic expression of these genes is plastic and strongly influenced by environmental factors. There was no evidence that local adaptation or outbreeding influenced otolith morphology or shape. Otoliths from fish exposed to thermal stress were bilaterally asymmetrical, whereas the bilateral symmetry of otoliths from outbred fish exhibited evidence of heterosis because they were more symmetrical than their native counterparts. Unlike development time and otoliths, gill raker development was linear and consistently stable in the face of both hybridization and environmental stress. These results make it clear that different biological attributes respond to genetic control and stress in different ways.
    • Genetic and phenotypic divergence within and between cinnamon teal (Anas cyanoptera) and blue-winged teal (A. discors)

      Wilson, Robert E.; McCracken, Kevin G.; Lindberg, Mark S.; Winker, Kevin; Sorenson, Michael D. (2011-08)
      Spatial heterogeneity in selection pressures can lead to extensive morphological variation and differences at functional genes between populations across a species' range without corresponding genetic variation at neutral loci. Divergent selection among populations may thus lead to intraspecific variation and in many cases speciation. Phenotypic and genetic structure within and between Cinnamon Teal (Anas cyanoptera) and the closely related Blue-winged Teal (A. discors) was assessed using body size measurements and neutral genetic markers in conjunction with a functional locus, hemoglobin. Cinnamon Teal are composed of five subspecies corresponding to distinct ecogeographic regions in North and South America. Subspecies and geographic regions differed significantly in overall body size, with the largest subspecies and the largest individuals found at high elevations in the central Andes (A. c. orinomus) and at high latitudes in southern Patagonia (A. c. cyanoptera). South American populations showed strong positive correlations with latitude and elevation while the migratory subspecies in North America (A. c. septentrionalium) showed few significant correlations with elevation and no relationship between latitude and body size. In addition, plumage differences were restricted to between North and South America as there was extensive variation observed within continents. Cinnamon Teal highland and lowland populations showed strong divergence in body size (Pst=0.56) and exhibited frequency differences in one non synonymous [alpha]-globin amino acid polymorphism (Asn/Ser-[alpha]9; Fst = 0.60), despite considerable admixture of reference loci. Selection pressures imposed by the hypoxic highland environment have likely resulted in asymmetric gene flow from the highlands into the lowlands following a highland colonization event from the lowlands. Cinnamon Teal and Blue-winged Teal show distinct but paraphyletic mitochondria) DNA ... and broadly shared nuclear alleles. Unlike South American Cinnamon Teal, North American Cinnamon Teal and Blue-winged Teal are characterized by high genetic diversity, large effective population size, and recent population expansion. Haplotypic and allelic sharing across continents is likely because of incomplete lineage sorting rather than ongoing gene flow. Within-continent estimates yielded higher migration rates consistent with hybridization. However, Cinnamon Teal and Blue-winged Teal are similar in body size; differences in plumage coloration may reduce hybridization events.
    • Genetic And Phenotypic Divergence Within And Between Cinnamon Teal (Anas Cyanoptera) And Blue-Winged Teal (A. Discors)

      Wilson, Robert E.; McCracken, Kevin; Sorenson, Michael D.; Winker, Kevin; Lindberg, Mark S. (2011)
      Spatial heterogeneity in selection pressures can lead to extensive morphological variation and differences at functional genes between populations across a species' range without corresponding genetic variation at neutral loci. Divergent selection among populations may thus lead to intraspecific variation and in many cases speciation. Phenotypic and genetic structure within and between Cinnamon Teal (Anas cyanoptera) and the closely related Blue-winged Teal (A. discors) was assessed using body size measurements and neutral genetic markers in conjunction with a functional locus, hemoglobin. Cinnamon Teal are composed of five subspecies corresponding to distinct ecogeographic regions in North and South America. Subspecies and geographic regions differed significantly in overall body size, with the largest subspecies and the largest individuals found at high elevations in the central Andes (A. c. orinomus) and at high latitudes in southern Patagonia (A. c. cyanoptera). South American populations showed strong positive correlations with latitude and elevation while the migratory subspecies in North America (A. c. septentrionalium) showed few significant correlations with elevation and no relationship between latitude and body size. In addition, plumage differences were restricted to between North and South America as there was extensive variation observed within continents. Cinnamon Teal highland and lowland populations showed strong divergence in body size (PST = 0.56) and exhibited frequency differences in one non-synonymous alpha-globin amino acid polymorphism (Asn/Ser-alpha9; FST = 0.60), despite considerable admixture of reference loci. Selection pressures imposed by the hypoxic highland environment have likely resulted in asymmetric gene flow from the highlands into the lowlands following a highland colonization event from the lowlands. Cinnamon Teal and Blue-winged Teal show distinct but paraphyletic mitochondrial DNA (phiST = 0.41) and broadly shared nuclear alleles. Unlike South American Cinnamon Teal, North American Cinnamon Teal and Blue-winged Teal are characterized by high genetic diversity, large effective population size, and recent population expansion. Haplotypic and allelic sharing across continents is likely because of incomplete lineage sorting rather than ongoing gene flow. Within-continent estimates yielded higher migration rates consistent with hybridization. However, Cinnamon Teal and Blue-winged Teal are similar in body size; differences in plumage coloration may reduce hybridization events.
    • Genetic consequences of ice ages for a holarctic rodent: phylogeography and post-glacial colonization of the tundra vole, Microtus oeconomus, in Beringia

      Galbreath, Kurt Egan (2002-08)
      Periodic glacial advances during the Pleistocene fragmented and displaced populations, while lowered sea levels permitted a biotic interchange between Asia and North America via the Bering Land Bridge. The tundra vole (Microtus oeconomus), a recent colonizer of North America, is a good model for studying the genetic consequences of these events. Variation in Mitochondrial (cytochrome b and control region) and nuclear (ALDH1) markers were examined within the context of Beringia's paleoclimatic history to examine the role of glaciations in driving differentiation and structuring patterns of genetic diversity. Genealogical relationships among genetic lineages were also assessed to elucidate probable paths of transberingian gene flow and post-glacial colonization. A deep phylogeographic break in western Beringia separates Beringian and Central Asian clades and may have been initiated by glacial vicariance. Population genetic structure within the Beringian clade has largely been determined by an historical reduction in genetic diversity and subsequent local differentiation. Serial bottlenecking during post-glacial colonization had minor effect, if any. Female-mediated gene flow among populations has been minimal since the last glacial maximum, but affinities among populations in Siberia and Alaska suggest two latitudinally partitioned routes of gene flow across the Bering Land Bridge. Also, post-glacial colonization of heavily glaciated southcoastal Alaska probably proceeded along coastal routes from the west after glacial recession.
    • Genetic diversity and population genetic structure of tanner crab Chionoecetes bairdi in Alaskan waters

      Johnson, Genevieve M.; López, J. Andrés; Eckert, Ginny L.; Hardy, Sarah M. (2019-05)
      Tanner crab (Chionoecetes bairdi) is a large-bodied species of crab harvested in commercial, personal use, and subsistence fisheries across Alaska. The commercial fisheries were highly productive until the 1980s, when most stocks faced major declines and were closed to harvest. The recovery success of stocks throughout the state has been variable throughout the subsequent decades, leading managers to question whether there are aspects of the population dynamics that are not accounted for. There is limited information on the genetic population structure of C. bairdi in Alaskan waters, which has caused uncertainty about whether established management areas align well with distribution and migration patterns for this species. I applied novel high throughput sequencing methods to measure genetic diversity and investigate the genetic population structure of C. bairdi in Alaskan waters. Genomic DNA was isolated from samples collected from Southeast Alaska, Prince William Sound, and the Eastern Bering Sea, both east and west of 166°W longitude, and processed according to a Double-Digest Restriction-Associated DNA Sequencing protocol. The final genotype assembly included 89 individuals that were genotyped at 2,740 independent, neutral single-nucleotide polymorphism (SNP) sites, and contained 3.06% missing data. The average observed heterozygosity across SNP sites within regions was significantly lower than the average heterozygosity expected for populations in Hardy-Weinberg equilibrium. An analysis of molecular variance indicated that genetic variability was mostly found within individuals (90%), 10% of variability was observed between individuals within sampling regions, and no significant amount of variation was detected between sampling regions. Furthermore, pairwise FST estimates between sampling regions were low, and thus the null model of panmixia could not be rejected. Principal components analysis was also congruent with a model of no differentiation among regions. Bayesian analysis implemented in the program STRUCTURE did not support any population partitioning above K = 1 clusters, again indicating that there is not substantial genetic differentiation among the regions sampled from across the state of Alaska. These results indicate high gene flow throughout the distribution of Tanner crab across the Alaska continental shelf. Recognized stocks are genetically indistinguishable from each other. This may indicate that stocks exchange a substantial number of migrants, and may not operate independently. This new information can provide insights as management plans are evaluated and refined.
    • Genetic engineering and characterization of LysR-type transcriptional regulators

      Sun, Honghong (2000-12)
      This thesis describes research aimed at understanding the structure and function of LysR-type transcriptional regulators. I studied two LysR-type proteins. One from the archaeon Methanococcus jannaschii, MJ-LysR. The other is from Burkholderia cepacia, DgdR. The MJ-LysR is the first putative LysR-type transcriptional regulator found in archaea. It is surprising that a prokaryotic transcriptional regulator is present in archaea, whose basal transcription machinery and RNA polymerase are more closely related to those of eukaryotes. To elucidate the structure and function of M-LysR protein, the gene was subcloned and expressed in E. coli. The gene product was isolated and purified by heat treatment and size exclusion chromatography. An in vitro binding assay showed that the purified protein bound to the intergenic region between the lysR gene and its upstream gene specifically and selectively. The results also showed that the protein maintained its binding activity even at 94C̊. The DNA footprinting data demonstrated a 30 bp protected region. Thus, this protein probably regulates expression of its own structural gene and perhaps the adjacent upstream gene. DgdR protein from Burkholderia cepacia had been previously characterized. The previous study showed that 2-methylalanine, the inducer for the DgdR regulated dgdA gene expression, but not D or L-alanine induced the conformational changes on DNA-protein complex. To further confirm this result, eleven amino acids with structures similar to 2-methylalainine were tested for their ability on affecting the binding of the DgdR protein to its operator site. Among these amino acids tested, only 2-methylalanine, 1-aminocyclopentane-1-carboxylic acid, S-2-aminobutanoic acid, RS-isovaline, and 2-trifluoromethyl-2-aminobutanoic acid generated the measurable band shifting. D- or L-norvaline, 2,2-diethyl glycine, and 2-trifluoromethylalanine did not cause any measurable change. It was concluded that both alkyl side chain size and hydrophobicity are important for the inducer recognition and binding in this protein. To solve the problem in DgdR protein purification caused by low solubility of this protein, a dgdR fusion gene to malE gene was constructed. This fusion gene provides a useful tool to further study and crystallize the DgdR protein.
    • Genetic linkage mapping of allozyme loci in even- and odd-year pink salmon (Oncorhynchus gorbuscha)

      Matsuoka, Makoto P.; Smoker, W. W. (1998)
      Genetic linkage maps of allozyme loci were constructed in even- and odd-year pink salmon (Oncorhynchus gorbuscha). The loci were mapped based on the results of gene-centromere (G-C) mapping and joint segregation analysis. For G-C mapping, 160 gynogenetic progeny families were produced, and 8,080 progeny from 74 families were analyzed using starch gel electrophoresis and histochemical stain techniques. G-C distances of 37 loci ranged from 0.5 cM at sMDH-A1* to 50 cM at sMDH-B2*. Eleven loci showed high G-C distances (>45 cM), indicating that one crossover on one chromosome arm is usual in pink salmon. Variation observed at sMDH-B1,2* in even-year families suggests that both of this loci is polymorphic and that there is possible inter-broodline chromosomal variation. Large variation was observed among families in G-C distance at several loci. Whether the variation was a reflection of difference in physical position, recombination rate, or some other factors needs clarification using a technique such as physical mapping with FISH, because this variation affects results of gene mapping based on recombination frequency. For joint segregation analysis, 320 biparental families were produced, and 13,068 progeny from 164 families were electrophoretically analyzed. Joint segregation was analyzed at over 200 locus pairs. Combined this with data from G-C mapping, 14 linkage groups involving 26 loci were constructed. The linkage maps contain eight classical linkage groups and four pseudolinkage groups. Two linkage groups found in pink salmon were conserved in widely divergent vertebrate species. Recombination frequency between linked loci were different between sexes, and it tends to be reduced in males in pink salmon. The order of loci, which probably duplicated in the recent tetraploidization event, in linkage groups I (sAAT-3 * &rarr; mAH-4*) and III (mAH-3* &rarr; sAAT-4*) was reversed. This is evidence of paracentric inversion during salmonid evolution after the duplication. Development of additional markers that are common (homologous) to many species will be necessary to examine syntenic stability and rearrangement over the evolutionary period.
    • Genetic variation in muskoxen (Ovibos moschatus)

      Fleischman, Claire L. (1986-05)
      Populations of Alaskan muskoxen (Ovibos moschatus) are derived from 34 animals transplanted from East Greenland in the 1930s. The possibility of a founder effect following this transplant was investigated. Muscle, liver and plasma samples from 87 Alaskan animals and 39 Greenlandic animals were analyzed using polyacrylamide and starch gel electrophoresis. A total of 38 enzyme and non-enzymatic protein systems, coded by 58 presumptive loci, was tested for activity; 28 loci were considered usable. One locus (Esterase-2) was polymorphic; the proportion of polymorphic loci was 0.036 (95% criterion). The mean heterozygosity per individual was 0.006 in the Greenlandic population and 0.011 in the Alaskan population. The allele frequencies at the Est-2 locus were similar in both populations. No heterozygote deficiency and no evidence of a founder effect were seen in the Alaskan population. This may be a consequence of the low level of allozymic variation seen in muskoxen in general.