transgene techniques

Ping-Hsien Chen, Yu-Ju Chen, Yi-Fen Chen, Yi-Chen Yeh, Kuo-Wei Chang, Ming-Chih Hou, Wen-Chuan Kuo
Changes in mucosal microvascular networks, called intraepithelial papillary capillary loops (IPCL), are an important key factor for diagnosing early-stage oral cancer in vivo. Nevertheless, there are a lack of tools to quantify these changes objectively. This is the first study to quantify the IPCL changes in vivo to differentiate benign or malignant oral lesions by the optical coherence tomography (OCT) technique. K14-EGFP-miR-211-GFP transgenic mice were inducted by 4-Nitroquinoline-1-oxide to produce oral carcinogenesis in different stages, including normal, premalignancy and cancer...
March 1, 2020: Biomedical Optics Express
Pasquale Molinaro, Silvia Natale, Angelo Serani, Lucrezia Calabrese, Agnese Secondo, Valentina Tedeschi, Valeria Valsecchi, Anna Pannaccione, Antonella Scorziello, Lucio Annunziato
Since the discovery of the three isoforms of the Na+ /Ca2+ exchanger, NCX1, NCX2 and NCX3 in 1990s, many studies have been devoted to identifying their specific roles in different tissues under several physiological or pathophysiological conditions. In particular, several seminal experimental works laid the foundation for better understanding gene and protein structures, tissue distribution, and regulatory functions of each antiporter isoform. On the other hand, despite the efforts in the development of specific compounds selectively targeting NCX1, NCX2 or NCX3 to test their physiological or pathophysiological roles, several drawbacks hampered the achievement of these goals...
March 2, 2020: Cell Calcium
Charles Romeo, Shih-Heng Chen, Eugenia Goulding, Lucas Van Gorder, Maura Schwartz, Mitzie Walker, Gregory Scott, Erica Scappini, Manas Ray, Negin P Martin
Gene delivery to fertilized eggs is often the first step in creation of transgenic animals, CRISPR knock-out, or early developmental studies. The zona pellucida, a hardened glycoprotein matrix surrounding the mammalian fertilized eggs, often complicates gene delivery by forming a barrier against transfection reagents and viruses. High efficiency techniques to perforate or penetrate the zona allow for access and gene delivery to fertilized eggs. However, these techniques often rely on highly skilled technologists, are costly, and require specialized equipment for micromanipulation, laser perforation, or electroporation...
March 17, 2020: Biochemical and Biophysical Research Communications
Jonathan Stefanowski, Alexander F Fiedler, Markus Köhler, Robert Günther, Wjatscheslaw Liublin, Martin Tschaikner, Ariana Rauch, David Reismann, Romano Matthys, Reto Nützi, Maria Gabriele Bixel, Ralf H Adams, Raluca A Niesner, Georg N Duda, Anja E Hauser
Bone healing involves the interplay of immune cells, mesenchymal cells, and vasculature over the time course of regeneration. Approaches to quantify the spatiotemporal aspects of bone healing at cellular resolution during long bone healing do not yet exist. Here, a novel technique termed Limbostomy is presented, which combines intravital microendoscopy with an osteotomy. This design allows a modular combination of an internal fixator plate with a gradient refractive index (GRIN) lens at various depths in the bone marrow and can be combined with a surgical osteotomy procedure...
March 20, 2020: Cytometry. Part A: the Journal of the International Society for Analytical Cytology
Francesco Pancaldi, Luisa M Trindade
The biomass demand to fuel a growing global bio-based economy is expected to tremendously increase over the next decades, and projections indicate that dedicated biomass crops will satisfy a large portion of it. The establishment of dedicated biomass crops raises huge concerns, as they can subtract land that is required for food production, undermining food security. In this context, perennial biomass crops suitable for cultivation on marginal lands (MALs) raise attraction, as they could supply biomass without competing for land with food supply...
2020: Frontiers in Plant Science
David Ding, Yichen Lu, Ruoyu Zhao, Xin Liu, Chawina De-Eknamkul, Chi Ren, Armaghan Mehrsa, Takaki Komiyama, Duygu Kuzum
OBJECTIVE: To investigate chronic durability of transparent graphene electrodes fabricated on polyethylene terephthalate (PET) and SU-8 substrates for chronic in vivo studies. METHODS: We perform systematic accelerated aging tests to understand the chronic reliability and failure modes of transparent graphene microelectrode arrays built on PET and SU-8 substrates. We employ graphene microelectrodes fabricated on PET substrate in chronic in vivo experiments with transgenic mice...
March 17, 2020: IEEE Transactions on Bio-medical Engineering
Seiji Shiozawa, Mayutaka Nakajima, Junko Okahara, Yoko Kuortaki, Fumihiko Kisa, Sho Yoshimatsu, Mari Nakamura, Ikuko Koya, Mika Yoshimura, Yohei Sasagawa, Itoshi Nikaido, Erika Sasaki, Hideyuki Okano
Mammalian pluripotent stem cells are thought to exist in two states: naïve and primed states. Generally, unlike those in rodents, pluripotent stem cells in primates including humans are regarded as being in the primed pluripotent state. Recently, several groups reported the existence of naïve pluripotent stem cells in humans. In this study, we report the conversion of primed state embryonic stem cells from common marmoset, a New World monkey, to the naïve state by using transgenes. The cells showed typical naïve state features including dome-like colony morphology, growth factor requirement, gene expression profile, X chromosome activation state, and energy metabolic status...
March 18, 2020: Stem Cells and Development
Lena Behrmann, Jasmin Wellbrock, Walter Fiedler
IntroductionMyeloid malignancies are caused by uncontrolled proliferation of neoplastic cells and lack of mature hematopoietic cells. Beside intrinsic genetic and epigenetic alterations within the neoplastic population, abnormal function of the bone marrow stroma promotes the neoplastic process. To overcome the supportive action of the microenvironment, recent research focuses on the development of targeted therapies, inhibiting the interaction of malignant cells and niche cells.Areas coveredThis review covers regulatory networks and potential druggable pathways within the hematopoietic stem cell niche...
March 18, 2020: Expert Opinion on Therapeutic Targets
Kiran S Khandagale, Rahul Chavhan, Altafhusain B Nadaf
2-acetyl-1-pyrroline (2AP) is a principal aroma compound in scented rice and a mutation in betaine aldehyde dehydrogenase 2 ( OsBADH2 ) is responsible aroma in scented rice. The present study was aimed at inducing 2AP production in non-scented indica rice cultivar IR-64 by silencing OsBADH2 via RNAi technique. A vector pBSK was used for the construction of RNAi cassette and pRI101ON as a binary vector. Agrobacterium (GV3101)-mediated transformation was done using embryogenic calli of IR-64. The resultant transgenic lines showed up to 14-fold reduction in expression of OsBADH2 gene and 50% inhibition in enzyme activity...
April 2020: 3 Biotech
Kunihiko Hashimoto, Takashi Kaito, Masayuki Furuya, Shigeto Seno, Daisuke Okuzaki, Junichi Kikuta, Hiroyuki Tsukazaki, Hideo Matsuda, Hideki Yoshikawa, Masaru Ishii
Bone morphogenetic protein (BMP)-2 plays a central role in bone-tissue engineering because of its potent bone-induction ability. However, the process of BMP-induced bone formation in vivo remains poorly elucidated. Here, we aimed to establish a method for intravital imaging of the entire process of BMP-2-induced ectopic bone formation. Using multicolor intravital imaging in transgenic mice, we visualized the spatiotemporal process of bone induction, including appearance and motility of osteoblasts and osteoclasts, angiogenesis, collagen-fiber formation, and bone-mineral deposition...
March 16, 2020: Scientific Reports
G A Gerashchenkov, N A Rozhnova, B R Kuluev, O Yu Kiryanova, G R Gumerova, A V Knyazev, Z R Vershinina, E V Mikhailova, D A Chemeris, R T Matniyazov, An Kh Baimiev, I M Gubaidullin, Al Kh Baimiev, A V Chemeris
CRISPR/Cas technology of genome editing is a powerful tool for making targeted changes in the DNA of various organisms, including plants. The choice of the precise nucleotide sequence (protospacer) in the gene to be edited is important in the design of guide RNA, which can be carried out by specialized software. We review and compare all the known on-line and off-line resources for guide RNA design, with special attention paid to tools capable of searching for off-target edits sites in plant genomes. The use of Cas12a may be preferable to Cas9...
January 2020: Molekuliarnaia Biologiia
Lena Rubi, Jean-Marc Fritschy
As the main input nucleus of the basal ganglia, the striatum plays a central role in planning, control, and execution of movement and motor skill learning. More than 90% of striatal neurons, so-called medium spiny neurons (MSN), are GABAergic projection neurons, innervating primarily the substantia nigra pars reticulata or the globus pallidus internus. Remaining are GABAergic and cholinergic interneurons, synchronizing and controlling striatal output by reciprocal connections with MSN. Besides prominent local cholinergic influence, striatal function is globally regulated by dopamine (DA) from the nigrostriatal pathway...
March 11, 2020: Journal of Neurophysiology
Dinushan Nesan, Hayley F Thornton, Laronna C Sewell, Deborah M Kurrasch
The hypothalamus is a key homeostatic brain region and the primary effector of neuroendocrine signaling. Recent studies show that early embryonic developmental disruption of this region can lead to neuroendocrine conditions later in life, suggesting that hypothalamic progenitors might be sensitive to exogenous challenges. To study the behavior of hypothalamic neural progenitors, we developed a novel dissection methodology to isolate murine hypothalamic neural stem and progenitor cells at the early timepoint of embryonic day (E) 12...
March 10, 2020: Endocrinology
Susmita Sarangi, Chiranjib Mandal, Sourav Dutta, Pranit Mukherjee, Raju Mondal, S P Jeevan Kumar, P Ray Choudhury, Vijay Pratap Singh, Durgesh Kumar Tripathi, Asit B Mandal
This data deals with the optimization of microprojectile bombardment particles for efficient genetic transformation in an indica rice involving AmSOD gene for development of salinity tolerant transgenic lines [1]. In this study, various parameters such as effect of genotypes, helium pressure, osmoticum, explants, flight distance, particle size, particle volume, vacuum, carrier DNA and stopping screen properties have been evaluated to determine their role in transformation of indica rice involving AmSOD gene for development of salinity tolerant Pusa Basmati 1 rice variety...
April 2020: Data in Brief
Maria J Lopez, Yashar Seyed-Razavi, Takefumi Yamaguchi, Gustavo Ortiz, Victor G Sendra, Deshea L Harris, Arsia Jamali, Pedram Hamrah
Multiphoton intravital microscopy (MP-IVM) is a powerful tool to image cells in vivo . Its application in immunology research has opened new horizons, allowing intravital imaging of leukocytes at the single-cell level. A transparent cornea is vital to retain vision. As an immune privileged site, a rapid innate response to foreign antigens is crucial in clearing opportunistic bacterial and viral pathogens, and minimizing collateral structural damage to the cornea. Furthermore, dissecting the mechanisms and preventing the immunological rejection process after corneal transplantation is imperative to retain sight...
2020: Frontiers in Immunology
Takashi Moriwaki, Satoshi Abe, Mitsuo Oshimura, Yasuhiro Kazuki
"Genomically" humanized animals are invaluable tools for generating human disease models and for biomedical research. Humanized animal models have generally been developed via conventional transgenic technologies; however, conventional gene delivery vectors such as viruses, plasmids, bacterial artificial chromosomes, P1 phase-derived artificial chromosomes, and yeast artificial chromosomes have limitations for transgenic animal creation as their loading gene capacity is restricted, and the expression of transgenes is unstable...
March 3, 2020: Experimental Cell Research
Kathleen E Grogan, George H Perry
In recent years, tools for functional genomic studies have become increasingly feasible for use by evolutionary anthropologists. In this review, we provide brief overviews of several exciting in vitro techniques that can be paired with "-omics" approaches (e.g., genomics, epigenomics, transcriptomics, proteomics, and metabolomics) for potentially powerful evolutionary insights. These in vitro techniques include ancestral protein resurrection, cell line experiments using primary, immortalized, and induced pluripotent stem cells, and CRISPR-Cas9 genetic manipulation...
March 6, 2020: Evolutionary Anthropology
Ashley L Cooney, Patrick L Sinn
Gene therapy for airway diseases requires efficient delivery of nucleic acids to the airways. In small animal models, gene delivery reagents are commonly delivered as a bolus dose. However, large animal models are often more relevant for the transition from preclinical studies to human trials. Aerosolizing viral vectors to the lungs of large animals can maximize anatomical distribution. Here, we describe a technique for aerosolization of viral vectors to the airways of newborn pigs. Briefly, a pig is anesthetized and intubated with an endotracheal tube, and a microsprayer is passed through the endotracheal tube...
March 6, 2020: BioTechniques
Victoire Cardot-Ruffino, Véronique Chauvet, Cassandre Caligaris, Adrien Bertrand-Chapel, Nicolas Chuvin, Roxane M Pommier, Ulrich Valcourt, David F Vincent, Sylvie Martel, Sophie Aires, Bastien Kaniewski, Pierre Dubus, Philippe Cassier, Stéphanie Sentis, Laurent Bartholin
Transforming growth factor (TGFβ) is a secreted factor, which accumulates in tissues during many physio- and pathological processes such as embryonic development, wound healing, fibrosis and cancer. In order to analyze the effects of increased microenvironmental TGFβ concentration in vivo, we developed a conditional transgenic mouse model (Flpo/Frt system) expressing bioactive TGFβ in fibroblasts, a cell population present in the microenvironment of almost all tissues. To achieve this, we created the genetically-engineered [Fsp1-Flpo; FSF TGFβCA ] mouse model...
March 3, 2020: Scientific Reports
Tania Janeth Porras-Gómez, Norma Moreno-Mendoza
It is widely accepted that the oocyte plays a very active role in promoting the growth of the follicle by directing the differentiation of granulosa cells and secreting paracrine growth factors. In turn, granulosa cells regulate the development of the oocytes, establishing close bidirectional communication between germ and somatic cells. The presence of cortical cells with morphological characteristics, similar to primordial germ cells that express specific germline markers, stem cells and cell proliferation, known as adult cortical germ cells (ACGC) have been reported in phyllostomid bats...
March 3, 2020: Zygote: the Biology of Gametes and Early Embryos
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