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Genome-wide association studies (GWAS) have become increasingly widely used to determine regions of the genome which may contain loci influencing the risk of neurological disorders. While linkage studies have identified genes that cause a number of Mendelian disorders, linkage analysis is less well suited for the more common complex disorders. This has led to the widespread use of GWAS for that purpose. Here we present and discuss several of the major extant GWAS in neurological disorders, their limitations, and implications of findings to date.

Recently, mutations in DCTN1were found to cause Perry syndrome, a parkinsonian disorder with TDP-43-positive pathology. Previously, mutations in DCTN1were identified in a family with lower motor neuron disease, in amyotrophic lateral sclerosis (ALS), and in a family with ALS/frontotemporal dementia (FTD), suggesting a central role for DCTN1in neurodegeneration.

One of the most frequently used tools in plant biotechnology, which includes genomics and proteomics, is gel electrophoresis. Our experience with middle and high school students as well as teachers and undergraduate students is that they have very little, if any, hands-on experience with this technique. These exercises were developed to demonstrate the principles of electrophoresis and DNA fingerprinting in middle and high school and university laboratories with minimal expense and equipment. The experiments have been tested by middle and high school students, as well as by teachers, and undergraduate and graduate students. The first exercise, electrophoresis of common food dyes, is primarily designed for secondary and undergraduate students, but can be used as an inexpensive means for introducing the main concepts of electrophoresis to anyone who has little or no experience, including graduate students. Popular brands of food dyes (red, blue, yellow, and green) purchased at local markets are mixed into a 60% glycerol/water solution and are separated on 1% agarose gels using 100 V for 35 min. Mixed colors are separated into primary colors (e.g., green into blue and yellow) and some apparently single dyes often have extra "surprise" components. A simple exercise illustrating forensic use of gel electrophoresis with dyes is also included. Over 100 students and teachers have completed this experiment successfully. The second laboratory exercise requires more extensive equipment and a more advanced set of skills; however, the exercise has been completed successfully by middle school -level through graduate-level students and by teachers. In this exercise, the internally transcribed spacer region of the ribosomal subunit for a fungus, plant, and insect are amplified and separated electrophoretically on agarose gels. A simple crime is solved using polymerase chain reaction (PCR) and DNA fingerprinting. The experiment protocol provides students with hands-on activities that include assembling master mixes for PCR, practice using pipettes, and performing the various steps involved in PCR amplification. Instructions for both exercises are formatted in easy-to -follow procedure boxes, and a downloadable presentation is available on the web. The cost of the expendables is about $1 per student, making these exercises relatively inexpensive to conduct, assuming that hardware and DNA are available. [ABSTRACT FROM AUTHOR]

The possibility of supplementing livestock diets with an aluminosilicate to protect them from fescue toxicosis was investigated. An in vitro study showed that hydrated sodium calcium aluminosilicate (HSCAS) removed > 90% of the ergotamine from aqueous solutions at pH 7.8 or lower, indicating a high affinity of ergotamine for HSCAS in vitro. Rats fed diets containing tall fescue seed infested (E+) with the endophytic fungus Acremonium coenophialum had lower (P< .05) feed intakes and weight gains than did rats fed diets containing uninfested (E-) tall fescue seed. When feed intake by rats fed the E- seed diet was limited to that of rats fed the E+ seed diet, weight gains did not differ, but testes weights and serum prolactin (PRL) concentrations were lower (P< .05 and .10, respectively) in rats receiving E+ seed. Supplementing E+ seed diets with HSCAS did not eliminate effects of E+ seed on intake, PRL, or testes weights. Sheep fed E+ tall fescue hay had higher (P< .05) rectal temperatures than did sheep fed an equal amount of E- tall fescue hay, but OM and N digestion coefficients did not differ between the two hays. Supplementing E+ hay diets with HSCAS did not eliminate the effect of E+ hay on rectal temperatures. Addition of 2% HSCAS to tall fescue hay diets did not affect apparent absorption by sheep of OM, N, Ca, P, Na, K, or Cu, but it reduced (P< .05) the apparent absorption of Mg, Mn, and Zn.(ABSTRACT TRUNCATED AT 250 WORDS)

Polyomavirus large T antigen binds to two sites located between positions -110 and -170 of a human heat shock protein 70 (hsp70) promoter. Methylation interference studies show that binding for each site is determined by two GPuGGC pentanucleotide sequences. The specificity of this binding interaction is similar to that observed for large T binding to the viral genome. The existence of sequences that bind a viral protein in a cellular promoter raises the possibility that these sequences play a role in gene expression in an uninfected cell. We show that hsp70 large T antigen binding site 1 is capable of functioning as an upstream promoter element in cells that do not contain any viral T antigen. Genetic analysis of this effect suggests that a cellular factor exists that has a binding specificity that overlaps but is not identical to that of polyomavirus large T antigen. To determine whether binding of polyomavirus large T antigen can regulate expression of the intact human hsp70 promoter, we have introduced the promoter into mouse cells with plasmids that express the polyomavirus early proteins. These proteins stimulate the level of correctly initiated hsp70 transcripts, but surprisingly the degree of stimulation remains unchanged for promoter constructs in which the large T antigen binding sites have been deleted. These observations suggest that trans activation of the hsp70 promoter by the polyomavirus early proteins occurs through protein-protein interactions and not through sequence-specific DNA binding.

A 3‐yr study was conducted at Ames Plantation, Grand Junction, TN to determine the effects of fungal endophyte (Acremonium conenophialumMorgan‐Jones & Gams) infestation (E+) levels in ‘Kentucky 31’ tall fescue (Festuca arundinaceaSchreb.) pastures on steer (Bos taurusL.) performance. Two replications were used of 1.2‐ha pastures overseeded yearly with ladino white clover (Trifolium repensL.) on a fine‐silty, mixed, thermic Typic Hapludalf soil. Yearling tester steers (three per pasture) and additional steers of similar size maintained available forage at heights of 2.5 to 7.5 an. Botanical composition, available forage, average daily gain (ADG), grazing days, and gain per hectare were determined every 21 d; serum prolactin, haircoat roughness, rectal temperatures, and E+levels were measured at longer intervals. Results were computed using a generalized linear mixed models procedure. Generalized least squares means and standard errors for the broad inference space were estimated (BLUE) and differences for specific linear contrasts were predicted (BLUP). Steer ADG was depressed from over 800 to about 450 g in spring and from about 700 to 400 g during the entire grazing season as A. coenophialumincreased from 3 to 81%. Animal grazing days were not different among treatments. Gain was about 65 kg ha−1more with 22% E+(379 kg ha−1) than at higher endophyte levels, but it was almost 500 kg ha−1at 3% E+. Serum prolactin (125 ng mL−1in steers grazing 3% E+) was halved with 22% E+and was depressed further at higher E+levels (41 to 18 ng mL−1). Results suggested that the effect of the endophyte on animal performance in tall fescue‐clover pastures is not linear, with greater depression per unit infestation at lower levels than at higher levels.

The human heat shock protein 70 (hsp70) gene is expressed constitutively in a wide variety of cells. Two separate promoter domains determine this basal level of hsp70 expression. The proximal domain is contained within 84 bases of the transcription initiation site and consists of three elements which appear to interact with the TATA factor(s) and CCAAT-box-binding transcription factor and SP1, respectively. The proximal domain is sufficient for near-maximal basal expression to rodent cell lines. The distal promoter domain consists of sequences upstream of -84 and is necessary in conjunction with the proximal domain for full basal expression in human cell lines. Although in BALB/c 3T3 cells the distal promoter domain plays little role in basal expression, it is functional as evidenced by the ability to compensate efficiently for mutations in the proximal CCAATC homology. The distal domain does not compensate as efficiently for proximal-domain mutations in HeLa cells. Basal expression of this human hsp70 promoter is, therefore, determined by multiple elements. Fewer elements are required for basal expression in rodent cell lines than in human cell lines, suggesting that there are significant differences between the rodent and human transcription apparatuses.

Abstract: Synthesis of ethylene in Kennebec potato tuber discs was measured at various time intervals after inoculation with zoospores of race 0 (incompatible) or race 1,2,3,4 (compatible) ofPhytophthora infestans (Mont.) de Bary. A burst of ethylene production was observed within 6 hr after inoculation with race 0, but not with race 1,2,3,4. This response may have been due to recognition of the incompatible race 0 during host penetration. Additional ethylene was synthesized concomitant with symptom appearance in both compatible and incompatible combinations.

We have made Chinese hamster ovary (CHO) cell lines that contain up to 2000 copies of the coding region of the mouse c-myc gene fused to the promoter of the Drosophila gene (hsp70) encoding a Mr 70,000 heat shock protein. Incubation of these cells at 43 degrees C results in an estimated 100-fold induction of c-myc mRNA. Translation of this mRNA occurs when the cells are returned to 37 degrees C, and during the first 3 hr of recovery at 37 degrees C, the c-myc protein is one of the most abundantly synthesized proteins in the cells. The products of the induced c-myc gene are phosphoproteins of apparent Mr 64,000, 66,000, and 75,000. Induced cells die, suggesting that elevated levels of c-myc are cytotoxic. Amplification of genes placed under control of the Drosophila hsp70 promoter may provide a general method for inducibly over expressing proteins in mammalian cells.

In vitro embryo‐derived callus culture is an efficient and useful technique for determining viable fungal endophyte (Acremonium coenophialumMorgan‐Jones & Gams) levels in tall fescue (Festuca arundinaceaSchreb.) seed lots; however, the presence of a yellow‐pigmented bacterium occasionally interferes with detection of the fungus. The objectives of this study were to control and identify this bacterium. Excised embryos from 26 seed lots were plated on Schenk and Hildebrandt medium amended with 20 μMdicamba (2‐methoxy‐3,6‐dichlorobenzoic acid); bacteria‐infested seeds were found in 22 lots. Infestation levels ranged from 1 to 100%. Medium supplemented with 50 mg L−1streptomycin eliminated bacterial growth without interfering with the growth of A. coenophialum. Based on microscopy, biochemical tests, and fatty acid analyses, the bacterium was identified as an Erwiniaspecies with characteristics of the herbicola group.