Bu araştırma, Tıp Araştırma Konseyi, Doğa Bilimleri ve Kanada&#39;nın Mühendislik Araştırma Konseyi ve Kanada İnme Ağ hibeler, tarafından desteklenmiştir. GAM Tıbbi Araştırma Alberta Heritage Vakfı tarafından desteklenmiştir.

Merdiven basamak yürüme test cihazı Metz ve Whishaw; yürüme test cihazı yatay merdiven basamak basamaklarının arasında 1 cm&#39;lik bir mesafe (bkz. Şekil 1 bir zemin oluşturmak için takılmış olabilir Pleksiglas ve metal basamaklara (3 mm çapında), yan duvarları , 2003). Ayrıca yan duvarlar yüksek, basamak yüksekliği ölçülür 1 m uzunluğunda ve 19 cm. Merdiven sonunda, nötr bir başlangıç ​​kafes ve bir sığınak (ev kafes) ile yerden 30 cm yükselmiş. Hayvanlar eğitim sırasında alışmış olduğu için, cihazın yükseklik anksiyete neden olma ihtimali oldu. Sokağın genişliği etrafında dönen hayvan önlemek için bir hayvan için yaklaşık 1 cm daha geniş, böylece hayvanın boyutuna göre ayarlandı. Görevin zorluk metal basamak konumunu değiştirerek güncellenmiştir. Basamaklarının düzenli bir desen çeşitli eğitim oturumları hayvanlar üzerinde desen öğrenmek ve basamaklarının pozisyonlar (Şekil 1, A Pattern) tahmin etmek için izin verdi. Deneme yargılanma değişti düzensiz bir desen desen (Şekil 1, Desen B) öğrenme hayvan engelledi. Basamaklara düzenli olarak düzenlenmesi için, 2 cm aralıklarla aralıklı idi. Düzensiz desen için basamak mesafe 1 ile 5 cm sistematik olarak değişiyordu. Beş düzensiz basamak kalıpları şablonları aynı desen (sonuçlar) test zorluk standartlaştırmak ve sonucu karşılaştırılabilirliği geliştirmek için tüm hayvanlar için uygulanmış ve böylece kullanılmıştır. İşitsel Bir kamera (Canovision, Canon Inc.), Hafif bir ventral açıyla yerleştirildi, böylece her dört bacağı pozisyonları aynı anda kayıt olabilir. Obtüratör hızı 500 - 2000 s Videorecordings 30 f / s hızla kare-kare analizi kullanılarak analiz edildi. Davranışsal eğitim ve test analizi Hayvanları evlerine kafes ulaşmak için nötr bir kafes merdiven geçmek için eğitilmiştir littermates evde kafes yürüyüş için pozitif pekiştirici. Bütün hayvanlar aynı yönde merdiven geçti. Merdiven geçmek için daha fazla takviye hayvanlar motive etmek için verildi. Bütün hayvanlar, eğitimli ve seans başına beş kez test edildi. Ayak hatası skorlama (Metz ve Whishaw, 2003) daha önce açıklanan bir ayak hatası skorlama sistemi kullanılarak, ön ayakları ve hindlimb yerleştirme nitel değerlendirme yapıldı. Analizi video kayıtlarını kare kare muayene yapıldı. Sadece her uzuv ardışık adımlar incelendi. Bu nedenle, böyle bir durak veya ayak hatası gibi bir yürüyüş kesintiye ve bir kesintiden sonra ilk adım önce son adımı attı. Merdivenin sonunda yapılan son adımlama döngüsü de puanlama çıkarılmıştır. Ekstremite yerleştirme bakımından bir özledim meydana basamaklarının arasında bir basamak ve uzuv çıkıntısı uzuv yerleştirme puan oldu. Tür basamaklara yürüyerek ya da pençe yerleştirme (bkz. Şekil 2) 7 kategori skalası kullanılarak değerlendirildi. Basamak yürüyerek ya da pençe yerleştirme konumlarını ve yerleştirme doğruluğu meydana gelen hatalar göre oylandı. (0) Toplam özledim. 0 puan uzuv tamamen bir basamak cevapsız verildi, yani dokunma vermedi ve bir düşme oluştu. Bir sonbaharda derin arasındaki basamaklara ve vücut duruş ve denge rahatsız düşen bir uzuv olarak tanımlanmıştır. (1) Derin kayma uzuv başlangıçta bir basamak üzerine yerleştirilir, sonra zaman ağırlık taşıyan kapalı terlik ve bir düşüş neden oldu. (2) hafif bir kayma uzuv bir basamak üzerinde yer, zaman ağırlık taşıyan kapalı terlik, ancak düşme sonucu ne yürüyüş döngüsü kesmedi. Bu durumda, hayvanın dengesini korumak ve koordineli bir yürüyüş devam başardı. (3) Yedek uzuv bir basamak üzerine yerleştirildi, ancak bu ağırlığı taşımaktan önce hızla kaldırdı ve başka bir basamak üzerine yerleştirilir. (4) Düzeltme. Uzuv bir basamak amaçlayan, ancak daha sonra ilk dokunmadan başka bir basamak üzerine yerleştirilmiştir. Alternatif olarak, bir uzuv, bir basamak üzerine yerleştirilen ise 4 puan kaydedildi ve aynı basamağı kalırken hızlı bir şekilde yeniden konumlandırılmış. (5) Kısmi yerleştirme bacak bilek ya da ön ayakları veya topuk veya hindlimb ayak parmakları iki basamak basamak yerleştirilmiştir. (6) doğru yerleştirilmesi bir bacak palmiye midportion tam ağırlığı desteği ile basamak yerleştirilmiştir. Farklı hataları aynı anda oluştuğu zaman, en düşük puan kaydedildi. Örneğin, bir ayak basamak ilk yerleştirilir ve daha sonra aynı adımı (skor 3) başka bir yer, ve daha sonra 1 puan süzüldü ve basamaklara (1 puan) arasında düştü ise kaydedildi. Whbir düşüş meydana geldi tr, sadece hata başlatan uzuv oylandı ve hayvan bütün uzuvları repositioned kadar diğer uzuvları hiçbiri gol oldu. Analizi için beş çalışmada hata puanları ortalamaları alındı. Ayak yerleştirme doğruluğu analizi (hata sayısı) Her geçerken hataların sayısı sayılmıştır. Hatalar ayak hatası skorlama sistemi göre belirlenmiştir. 0, 1 veya 2 puan bir puan alan her uzuv yerleşim olarak tanımlanan bir hata, bir hata, ayağı kayma veya toplam özledim her türlü temsil eder yani. Hataları ve adım sayısı, her bacak için ayrı ayrı kaydedildi. Bu verilere göre, adım başına hataların ortalama sayısı hesaplanmış ve beş denemeleri için ortalama. Analiz ikinci nicel parametre merdiven görevi tüm uzunluğu geçmek için gerekli ortalama süre oldu. Zaman ölçümü hayvan merdiven yerleştirildi sonra başlayan ve tüm uzunluğu yürümeye başladı. Bir hayvan, bir stop harcanan zaman ölçüm dahil değildi. Forepaw haneli puan Forepaw haneli skor pençe, bir basamak üzerindeki midportion (yani, ayak hatası skorlama sistemi skor 6) ile doğru yerleştirildi kaydedildi. Forepaw rakamlar etrafında fleksiyon bir basamak olabilir derecesi (Şekil 2), üç noktalı bir ölçekte oylandı. Puan ön ayakları bir basamak üzerinde dikey olarak konumlandırılmış, yani maksimum ağırlık desteği ile pozisyon uygulandı. Puanlar şöyle verildi: (0) Rakamlar yaklaşık 90 derecelik bir açı kapattı. (1) Rakamlar yaklaşık 45 derecelik bir açı kapattı. (2) Rakamlar tamamen basamak çevresinde fleksiyondayken. Beş adım puanları ortalaması ve analizi için kullanılır. Şekil açıklamaları <img src="/files/ftp_upload/old/1204_2009_1_20_6/Fig 1 Grid.png" alt="Şekil 1" /> Şekil 1 frontolateral görünümünde test aparatı yürüyüş yetenekli cihazın ölçülerini, merdiven basamak belirtti. Bu çalışmada iki farklı basamak desen kullanılmıştır: Desen, düzenli basamak düzenleme, Desen B, düzensiz basamak düzenleme. Düzensiz basamak düzenleme ardışık denemeler arasında değişmektedir. <img src="/files/ftp_upload/old/1204_2009_1_20_7/Fig 2 (angle lines).png" alt="Şekil 2" /> Şekil 2. Temsilcisi fotoğraflar haneli skor üç kategoride gösteren. Tam ağırlık desteği ön ayakları ile ilgili basamak açısı Not.

<ol>
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Q., Gorny, B., Sarna, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Paw+and+limb+use+in+skilled+and+spontaneous+reaching+after+pyramidal+tract,+red+nucleus+and+combined+lesions+in+the+rat:+behavioral+and+anatomical+dissociations.">Paw and limb use in skilled and spontaneous reaching after pyramidal tract, red nucleus and combined lesions in the rat: behavioral and anatomical dissociations.</a> Behav Brain Res. 93, 167-183 (1998).</li><li>Z'Graggen, W. J., Metz, G. A. S., Kartje, G. L., Schwab, M. 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Bu çalışmada, yetenekli yürüyüş, bacak yerleştirme ve bacak koordinasyonu değerlendirmek için yeni bir test olarak görev yürüyen merdiven basamak tanıttı. Görev yürüyen merdiven basamak nitelikli yürüme kalitatif ve kantitatif analiz birleştirerek ince motor fonksiyon bozuklukları arasında ayırım sağlar. Bozuklukların daha açıklayıcı bir hesap için kullanılan video analiz prosedürü ve bir basamak açgözlü Bu yazıda anlatılan basit bir derecelendirme ölçeği ekstremite yerleşimi ...

the ladder rung walking task: a scoring system and its practical application.

/ Inme, omurilik yaralanması, ve diğer nörodejeneratif hastalık hayvan modellerinin geliştirilmesinde ilerleme, motor fonksiyon farklı yönlerini ayrıntılı ve hareket kapasitesinin bile ince kaybını belirlemek için, yüksek hassasiyet testleri gerektirir. Test etkinliği ve çözünürlüğü artırmak için, testleri, motor fonksiyon kalitatif ve kantitatif tedbirler izni ve iyileşme dönemlerinde performans değişikliklerine duyarlı olmalıdır. Bu çalışmada sıçan yetenekli yürüme değerlendirmek için aynı zamanda ön ayakları ve hindlimb fonksiyonu ölçmek için yeni bir görev açıklamaktadır. Hayvanlar basamak boşluğu değişkendir ve periyodik olarak değiştirilmelidir üzerinde yatay bir merdiven boyunca yürümek için gereklidir. Basamak aralık değişiklikler basamaklarının mutlak ve göreli konumunu öğrenme hayvanlar önlemek ve böylece, hayvanların öğrenme yoluyla bozuklukları telafi etmek için yeteneği en aza indirmek. Buna ek olarak, basamaklara arasındaki aralığı değiştirerek testi, uzun vadeli çalışmalar tekrar tekrar kullanılmasına imkan verir. Yöntemleri uzuv yerleştirerek, adım, ortak koordinasyon da dahil olmak üzere, ön ve hindlimb performans hem nicel hem de nitel bir açıklama için açıklanmıştır. Ayrıca, telafi edici stratejileri kullanımı missteps veya başka bir uzuv yanlış yerleştirilmesini telafi edici adımlar tarafından gösterilir.

Watch this Scientific Journal Video about The Ladder Rung Walking Task: A Scoring System and its Practical Application. at JoVE.com

The Ladder Rung Walking Task: A Scoring System and its Practical Application.

Görev yürüyen merdiven basamak yetenekli yürüme değerlendirmek ve ön ayakları ve hindlimb, yerleştirme adım ve ekstremite arası koordinasyon ölçmek için yeni bir testtir.

Görev Yürüyüş Merdiven Basamak: Puanlama Sistemi ve Pratik Uygulama.

Progress in the development of animal models for/stroke, spinal cord injury, and other neurodegenerative disease requires tests of high sensitivity to ...

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25.9K Görüntüleme.  University of Lethbridge. Görev yürüyen merdiven basamak yetenekli yürüme değerlendirmek ve ön ayakları ve hindlimb, yerleştirme adım ve ekstremite arası koordinasyon ölçmek için yeni bir testtir.

Video: The Ladder Rung Walking Task: A Scoring System and its Practical Application.

Induction and Clinical Scoring of Chronic-Relapsing Experimental Autoimmune Encephalomyelitis

Multiple sclerosis (MS) is a chronic inflammatory disease of the central nervous system (CNS) that commonly affects young adults. It is characterized by demyelination and glial scaring in areas disseminated in the brain and spinal cord. These lesions alter nerve conduction and induce the disabling neurological deficits that vary with the location of the demyelinated plaques in the CNS (e.g. paraparesis, paralysis, blindness, incontinence).
Experimental autoimmune encephalomyelitis (EAE) is a model for MS. EAE was first induced accidentally in humans during vaccination against rabies, using viruses grown on rabbit spinal cords. Residues of spinal injected with the inactivated virus induced the CNS disease. Following these observations, a first model of EAE was described in non-human primates immunized with a CNS homogenate by Rivers and Schwenther in 1935. EAE has since been generated in a variety of species and can follow different courses depending on the species/strain and immunizing antigen used. For example, immunizing Lewis rats with myelin basic protein in emulsion with adjuvant induces an acute model of EAE, while the same antigen induces a chronic disease in guinea pigs.
The EAE model described here is induced by immunizing DA rats against DA rat spinal cord in emulsion in complete Freund's adjuvant. Rats develop an ascending flaccid paralysis within 7-14 days post-immunization. Clinical signs follow a relapsing-remitting course over several weeks. Pathology shows large immune infiltrates in the CNS and demyelination plaques. Special considerations for taking care for animals with EAE are described at the end of the video.

This video demonstrates the induction and clinical scoring of an animal model of multiple sclerosis: chronic-relapsing experimental autoimmune encephalomyelitis in DA rats. The disease, induced by immunizing rats with an emulsion containing whole rat spinal cord and complete Freund's adjuvant, presents clinical signs resembling the human disease.

İndüksiyon ve Kronik Relaps Deneysel Otoimmün Encephalomyelitis Klinik Puanlama

Multiple sclerosis (MS) is a chronic inflammatory disease of the central nervous system (CNS) that commonly affects young adults. It is characterized by ...

Biyoloji

Use of Rotorod as a Method for the Qualitative Analysis of Walking in Rat

High speed videoanalysis of the details of movement can provide a source of information about qualitative aspects of walking movements. When walking on a rotorod, animals remain in approximately the same place making repetitive movements of stepping. Thus the task provides a rich source of information on the details of foot stepping movements. Subjects were hemi-Parkinson analogue rats, produced by injection of 6-hydroxydopamine (6-OHDA) into the right nigrostriatal bundle to deplete nigrostriatal dopamine (DA). The present report provides a video analysis illustration of animals previously were filmed from frontal, lateral, and posterior views as they walked (15). Rating scales and frame-by-frame replay of the video records of stepping behavior indicated that the hemi-Parkinson rats were chronically impaired in posture and limb use contralateral to the DA-depletion. The contralateral limbs participated less in initiating and sustaining propulsion than the ipsilateral limbs. These deficits secondary to unilateral DA-depletion show that the rotorod provides a use task for the analysis of stepping movements.

The rotorod test is used to assess motor status in the walking movement of hemi-Parkinson analogue rats.

Rotorod Rat Yürüyüş Kalitatif Analiz için bir yöntem olarak kullanımı

High speed videoanalysis of the details of movement can provide a source of information about qualitative aspects of walking movements. When walking on a ...

Lens Transplantation in Zebrafish and its Application in the Analysis of Eye Mutants

The lens plays an important role in the development of the optic cup[1,2]. Using the zebrafish as a model organism, questions regarding lens development can be addressed. The zebrafish is useful for genetic studies due to several advantageous characteristics, including small size, high fecundity, short lifecycle, and ease of care. Lens development occurs rapidly in zebrafish. By 72 hpf, the zebrafish lens is functionally mature [3]. Abundant genetic and molecular resources are available to support research in zebrafish. In addition, the similarity of the zebrafish eye to those of other vertebrates provides basis for its use as an excellent animal model of human defects[4-7]. Several zebrafish mutants exhibit lens abnormalities, including high levels of cell death, which in some cases leads to a complete degeneration of lens tissues [8]. To determine whether lens abnormalities are due to intrinsic causes or to defective interactions with the surrounding tissues, transplantation of a mutant lens into a wild-type eye is performed. Using fire-polished metal needles, mutant or wild-type lenses are carefully dissected from the donor animal, and transferred into the host. To distinguish wild-type and mutant tissues, a transgenic line is used as the donor. This line expresses membrane-bound GFP in all tissues, including the lens. This transplantation technique is an essential tool in the studies of zebrafish lens mutants.

Lens development involves interactions with other tissues. Several zebrafish eye mutants are characterized by an abnormally small lens size. Here we demonstrate a lens transplantation experiment to determine whether this phenotype is due to intrinsic causes or defective interactions with tissues that surround the lens.

Zebra balığı Lens Nakli ve Göz Mutants Analizi Uygulaması

The lens plays an important role in the development of the optic cup[1,2]. Using the zebrafish as a model organism, questions regarding lens development ...

Tomato Analyzer: A Useful Software Application to Collect Accurate and Detailed Morphological and Colorimetric Data from Two-dimensional Objects

Measuring fruit morphology and color traits of vegetable and fruit crops in an objective and reproducible way is important for detailed phenotypic analyses of these traits. Tomato Analyzer (TA) is a software program that measures 37 attributes related to two-dimensional shape in a semi-automatic and reproducible manner1,2. Many of these attributes, such as angles at the distal and proximal ends of the fruit and areas of indentation, are difficult to quantify manually. The attributes are organized in ten categories within the software: Basic Measurement, Fruit Shape Index, Blockiness, Homogeneity, Proximal Fruit End Shape, Distal Fruit End Shape, Asymmetry, Internal Eccentricity, Latitudinal Section and Morphometrics. The last category requires neither prior knowledge nor predetermined notions of the shape attributes, so morphometric analysis offers an unbiased option that may be better adapted to high-throughput analyses than attribute analysis. TA also offers the Color Test application that was designed to collect color measurements from scanned images and allow scanning devices to be calibrated using color standards3. 
TA provides several options to export and analyze shape attribute, morphometric, and color data. The data may be exported to an excel file in batch mode (more than 100 images at one time) or exported as individual images. The user can choose between output that displays the average for each attribute for the objects in each image (including standard deviation), or an output that displays the attribute values for each object on the image. TA has been a valuable and effective tool for indentifying and confirming tomato fruit shape Quantitative Trait Loci (QTL), as well as performing in-depth analyses of the effect of key fruit shape genes on plant morphology. Also, TA can be used to objectively classify fruit into various shape categories. Lastly, fruit shape and color traits in other plant species as well as other plant organs such as leaves and seeds can be evaluated with TA.

Tomato Analyzer (TA) quantifies attributes of two dimensional shapes and color in a reproducible and accurate manner. A step-by-step procedure for obtaining high quality digitalized images of tomato fruit, morphological and color analyses of these images and several applications using the data generated through this software are described.

Domates Analyzer: İki boyutlu Objects Doğru ve detaylı Morfolojik ve Kolorimetrik Veri Toplama Faydalı Yazılım Uygulaması

Measuring fruit morphology and color traits of vegetable and fruit crops in an objective and reproducible way is important for detailed phenotypic ...

Visualization of Caenorhabditis elegans Cuticular Structures Using the Lipophilic Vital Dye DiI

The cuticle of C. elegans is a highly resistant structure that surrounds the exterior of the animal1-4. The cuticle not only protects the animal from the environment, but also determines body shape and plays a role in motility4-6. Several layers secreted by epidermal cells comprise the cuticle, including an outermost lipid layer7.
Circumferential ridges in the cuticle called annuli pattern the length of the animal and are present during all stages of development8. Alae are longitudinal ridges that are present during specific stages of development, including L1, dauer, and adult stages2,9. Mutations in genes that affect cuticular collagen organization can alter cuticular structure and animal body morphology5,6,10,11. While cuticular imaging using compound microscopy with DIC optics is possible, current methods that highlight cuticular structures include fluorescent transgene expression12, antibody staining13, and electron microscopy1. Labeled wheat germ agglutinin (WGA) has also been used to visualize cuticular glycoproteins, but is limited in resolving finer cuticular structures14. Staining of cuticular surface using fluorescent dye has been observed, but never characterized in detail15. We present a method to visualize cuticle in live C. elegans using the red fluorescent lipophilic dye DiI (1,1'-dioctadecyl-3,3,3',3'-tetramethylindocarbocyanine perchlorate), which is commonly used in C. elegans to visualize environmentally exposed neurons. This optimized protocol for DiI staining is a simple, robust method for high resolution fluorescent visualization of annuli, alae, vulva, male tail, and hermaphrodite tail spike in C. elegans.

Visualization of Caenorhabditis elegans Cuticular Structures Using the Lipophilic Vital Dye DiI

We present a method to visualize cuticle in live C. elegans using the red fluorescent lipophilic dye DiI (1,1'-dioctadecyl-3,3,3',3'-tetramethylindocarbocyanine perchlorate), which is commonly used in C. elegans to visualize environmentally exposed neurons. With this optimized protocol, alae and annular cuticular structures are stained by DiI and observed using compound microscopy.

Görselleştirme Caenorhabditis Elegans Tırnak Yapıları Lipofilik Vital Boya, DII

The cuticle of C. elegans is a highly resistant structure that surrounds the exterior of the animal1-4. The cuticle not only protects the animal from the ...

RNAi Screening to Identify Postembryonic Phenotypes in C. elegans

C. elegans has proven to be a valuable model system for the discovery and functional characterization of many genes and gene pathways1. More sophisticated tools and resources for studies in this system are facilitating continued discovery of genes with more subtle phenotypes or roles. 
Here we present a generalized protocol we adapted for identifying C. elegans genes with postembryonic phenotypes of interest using RNAi2. This procedure is easily modified to assay the phenotype of choice, whether by light or fluorescence optics on a dissecting or compound microscope. This screening protocol capitalizes on the physical assets of the organism and molecular tools the C. elegans research community has produced. As an example, we demonstrate the use of an integrated transgene that expresses a fluorescent product in an RNAi screen to identify genes required for the normal localization of this product in late stage larvae and adults. First, we used a commercially available genomic RNAi library with full-length cDNA inserts. This library facilitates the rapid identification of multiple candidates by RNAi reduction of the candidate gene product. Second, we generated an integrated transgene that expresses our fluorecently tagged protein of interest in an RNAi-sensitive background. Third, by exposing hatched animals to RNAi, this screen permits identification of gene products that have a vital embryonic role that would otherwise mask a post-embryonic role in regulating the protein of interest. Lastly, this screen uses a compound microscope equipped for single cell resolution.

RNAi Screening to Identify Postembryonic Phenotypes in C. elegans

We describe a sensitized method to identify postembryonic regulators of protein expression and localization in C. elegans using an RNAi-based genomic screen and an integrated transgene that expresses a functional, fluorescently tagged protein.

Içinde Postembryonic Fenotiplerin Belirleme RNAi Tarama C. elegans</em

C. elegans has proven to be a valuable model system for the discovery and functional characterization of many genes and gene pathways1. More sophisticated ...

A Fluorescence-based Exonuclease Assay to Characterize DmWRNexo, Orthologue of Human Progeroid WRN Exonuclease, and Its Application to Other Nucleases

WRN exonuclease is involved in resolving DNA damage that occurs either during DNA replication or following exposure to endogenous or exogenous genotoxins. It is likely to play a role in preventing accumulation of recombinogenic intermediates that would otherwise accumulate at transiently stalled replication forks, consistent with a hyper-recombinant phenotype of cells lacking WRN. In humans, the exonuclease domain comprises an N-terminal portion of a much larger protein that also possesses helicase activity, together with additional sites important for DNA and protein interaction. By contrast, in Drosophila, the exonuclease activity of WRN (DmWRNexo) is encoded by a distinct genetic locus from the presumptive helicase, allowing biochemical (and genetic) dissection of the role of the exonuclease activity in genome stability mechanisms. Here, we demonstrate a fluorescent method to determine WRN exonuclease activity using purified recombinant DmWRNexo and end-labeled fluorescent oligonucleotides. This system allows greater reproducibility than radioactive assays as the substrate oligonucleotides remain stable for months, and provides a safer and relatively rapid method for detailed analysis of nuclease activity, permitting determination of nuclease polarity, processivity, and substrate preferences.

Exonucleases play critical roles in ensuring genome stability. Loss of WRN exonuclease function results in premature aging. Studying substrates and other requirements of the nuclease in vitro can help elucidate its role in vivo. Here we demonstrate a rapid and reproducible fluorescence-based assay to measure its nuclease activity.

WRN exonuclease is involved in resolving DNA damage that occurs either during DNA replication or following exposure to endogenous or exogenous genotoxins. ...

A Practical Guide to Phylogenetics for Nonexperts

Many researchers, across incredibly diverse foci, are applying phylogenetics to their research question(s). However, many researchers are new to this topic and so it presents inherent problems. Here we compile a practical introduction to phylogenetics for nonexperts.&#160;We outline in a step-by-step manner, a pipeline for generating reliable phylogenies from gene sequence datasets. We begin with a user-guide for similarity search tools via online interfaces as well as local executables. Next, we explore programs for generating multiple sequence alignments followed by protocols for using software to determine best-fit models of evolution. We then outline protocols for reconstructing phylogenetic relationships via maximum likelihood and Bayesian criteria&#160;and finally describe tools for visualizing phylogenetic trees. While this is not by any means an exhaustive description of phylogenetic approaches, it does provide the reader with practical starting information on key software applications commonly utilized by phylogeneticists. The vision for this article would be that it could serve as a practical training tool for researchers embarking on phylogenetic studies&#160;and also serve as an educational resource that could be incorporated into a classroom or teaching-lab.

Here we describe a step-by-step pipeline for generating reliable phylogenies from nucleotide or amino acid sequence datasets. This guide aims to serve researchers or students new to phylogenetic analysis.&#160;

Nonexperts için Phylogenetics İçin Pratik Kılavuz

Many researchers, across incredibly diverse foci, are applying phylogenetics to their research question(s). However, many researchers are new to this ...

Design and Analysis of Temperature Preference Behavior and its Circadian Rhythm in Drosophila

The circadian clock regulates many aspects of life, including sleep, locomotor activity, and body temperature (BTR) rhythms1,2. We recently identified a novel Drosophila circadian output, called the temperature preference rhythm (TPR), in which the preferred temperature in flies rises during the day and falls during the night 3. Surprisingly, the TPR and locomotor activity are controlled through distinct circadian neurons3. Drosophila locomotor activity is a well&#160;known circadian behavioral output and has provided strong contributions to the discovery of many conserved mammalian circadian clock genes and mechanisms4. Therefore, understanding TPR will lead to the identification of hitherto unknown molecular and cellular circadian mechanisms. Here, we describe how to perform and analyze the TPR assay. This technique not only allows for dissecting the molecular and neural mechanisms of TPR, but also provides new insights into the fundamental mechanisms of the brain functions that integrate different environmental signals and regulate animal behaviors. Furthermore, our recently published data suggest that the fly TPR shares features with the mammalian BTR3. Drosophila are ectotherms, in which the body temperature is typically behaviorally regulated. Therefore, TPR is a strategy used to generate a rhythmic body temperature in these flies5-8. We believe that further exploration of Drosophila TPR will facilitate the characterization of the mechanisms underlying body temperature control in animals.

Design and Analysis of Temperature Preference Behavior and its Circadian Rhythm in Drosophila

We recently identified a novel Drosophila circadian output, temperature preference rhythm (TPR), in which the preferred temperature in flies rises during the day and falls during the night. TPR is regulated independently from another circadian output, locomotor activity. Here&#160;we describe the design and analysis of TPR in Drosophila.

Drosophila'da Sıcaklık Tercih Davranışının ve Sirkadiyen Ritminin Tasarımı ve Analizi

The circadian clock regulates many aspects of life, including sleep, locomotor activity, and body temperature (BTR) rhythms1,2. We recently identified a ...

Reconstruction of 3-Dimensional Histology Volume and its Application to Study Mouse Mammary Glands

Histology volume reconstruction facilitates the study of 3D shape and volume change of an organ at the level of macrostructures made up of cells. It can also be used to investigate and validate novel techniques and algorithms in volumetric medical imaging and therapies. Creating 3D high-resolution atlases of different organs1,2,3 is another application of histology volume reconstruction. This provides a resource for investigating tissue structures and the spatial relationship between various cellular features. We present an image registration approach for histology volume reconstruction, which uses a set of optical blockface images. The reconstructed histology volume represents a reliable shape of the processed specimen with no propagated post-processing registration error. The Hematoxylin and Eosin (H&#38;E) stained sections of two mouse mammary glands were registered to their corresponding blockface images using boundary points extracted from the edges of the specimen in histology and blockface images. The accuracy of the registration was visually evaluated. The alignment of the macrostructures of the mammary glands was also visually assessed at high resolution.
This study delineates the different steps of this image registration pipeline, ranging from excision of the mammary gland through to 3D histology volume reconstruction. While 2D histology images reveal the structural differences between pairs of sections, 3D histology volume provides the ability to visualize the differences in shape and volume of the mammary glands.

We present an image registration approach for 3-dimensional (3D) histology volume reconstruction, which facilitates the study of the changes of an organ at the level of macrostructures made up of cells . Using this approach, we studied the 3D changes between wild-type and Igfbp7-null mammary glands.

3-Boyutlu Histoloji hacmi ve Fare meme bezlerinin Eğitim için onun Uygulama İmar

Histology volume reconstruction facilitates the study of 3D shape and volume change of an organ at the level of macrostructures made up of cells. It can ...