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Transgenic Research

Kristin M Whitworth, Joshua A Benne, Lee D Spate, Stephanie L Murphy, Melissa S Samuel, Clifton N Murphy, Jürgen A Richt, Eric Walters, Randall S Prather, Kevin D Wells
The CRISPR/Cas9 genome editing tool has increased the efficiency of creating genetically modified pigs for use as biomedical or agricultural models. The objectives were to determine if DNA editing resulted in a delay in development to the blastocyst stage or in a skewing of the sex ratio. Six DNA templates (gBlocks) that were designed to express guide RNAs that target the transmembrane protease, serine S1, member 2 (TMPRSS2) gene were in vitro transcribed. Pairs of CRISPR guide RNAs that flanked the start codon and polyadenylated Cas9 were co-injected into the cytoplasm of zygotes and cultured in vitro to the blastocyst stage...
October 15, 2016: Transgenic Research
Ana Cecilia Gomes Silva, Daniela Volcan Almeida, Bruna Felix Nornberg, Jessica Ribeiro Pereira, Diego Martins Pires, Carine Dahl Corcini, Antonio Sergio Varela Junior, Luis Fernando Marins
Growth hormone (GH) transgenesis presents a high potential application in aquaculture. However, excess GH may have serious consequences due to pleiotropic actions. In order to study these effects in zebrafish (Danio rerio), two transgenic lines were developed. The first expresses GH ubiquitously and constitutively (F0104 line), while the second expresses the GH receptor in a muscle-specific manner (Myo-GHR line). Results from the F0104 line showed accelerated growth but increased reproductive difficulties, while Myo-GHR did not show the expected increase in muscle mass...
October 14, 2016: Transgenic Research
Jordi Ruiz-Camp, José Alberto Rodríguez-Castillo, Susanne Herold, Konstantin Mayer, István Vadász, Michelle D Tallquist, Werner Seeger, Katrin Ahlbrecht, Rory E Morty
Bronchopulmonary dysplasia (BPD) is the most common complication of preterm birth characterized by blunted post-natal lung development. BPD can be modelled in mice by exposure of newborn mouse pups to elevated oxygen levels. Little is known about the mechanisms of perturbed lung development associated with BPD. The advent of transgenic mice, where genetic rearrangements can be induced in particular cell-types at particular time-points during organogenesis, have great potential to explore the pathogenic mechanisms at play during arrested lung development...
October 11, 2016: Transgenic Research
Fei Xu, Xiuhua Kang, Liang Chen, Chuanhui Chen, Gen Hu, Wei Bai, Wei Zhang
Asthma is a common chronic inflammatory disease in the airways with wide prevalence, and it is thought to be caused by the combinational factors in environment and genetics. A large body of studies has suggested that cell immunity played a vital role in regulating the airway hyperreactivity (AHR) and inflammation. Therefore, we here developed a mouse model of asthma by microinjecting the pronucleus with a vector spontaneously coding human IL10 and TGFB1 gene to explore the possible interaction between these two potent molecules during asthma progression...
October 1, 2016: Transgenic Research
Nabeela Abid, Asia Khatoon, Asma Maqbool, Muhammad Irfan, Aftab Bashir, Irsa Asif, Muhammad Shahid, Asma Saeed, Henrik Brinch-Pedersen, Kauser A Malik
Phytate is a major constituent of wheat seeds and chelates metal ions, thus reducing their bioavailability and so the nutritional value of grains. Transgenic plants expressing heterologous phytase are expected to enhance degradation of phytic acid stored in seeds and are proposed to increase the in vitro bioavailability of mineral nutrients. Wheat transgenic plants expressing Aspergillus japonicus phytase gene (phyA) in wheat endosperm were developed till T3 generation. The transgenic lines exhibited 18-99 % increase in phytase activity and 12-76 % reduction of phytic acid content in seeds...
September 29, 2016: Transgenic Research
Xiaoli Xu, Cheng Peng, Xiaofu Wang, Xiaoyun Chen, Qiang Wang, Junfeng Xu
This study evaluated the applicability of droplet digital PCR (ddPCR) as a tool for maize zygosity determination using quantitative real-time PCR (qPCR) as a reference technology. Quantitative real-time PCR is commonly used to determine transgene copy number or GMO zygosity characterization. However, its effectiveness is based on identical reaction efficiencies for the transgene and the endogenous reference gene. Additionally, a calibrator sample should be utilized for accuracy. Droplet digital PCR is a DNA molecule counting technique that directly counts the absolute number of target and reference DNA molecules in a sample, independent of assay efficiency or external calibrators...
September 8, 2016: Transgenic Research
Jagdeep Kaur, John Fellers, Alok Adholeya, Siva L S Velivelli, Kaoutar El-Mounadi, Natalya Nersesian, Thomas Clemente, Dilip Shah
Rust fungi of the order Pucciniales are destructive pathogens of wheat worldwide. Leaf rust caused by the obligate, biotrophic basidiomycete fungus Puccinia triticina (Pt) is an economically important disease capable of causing up to 50 % yield losses. Historically, resistant wheat cultivars have been used to control leaf rust, but genetic resistance is ephemeral and breaks down with the emergence of new virulent Pt races. There is a need to develop alternative measures for control of leaf rust in wheat. Development of transgenic wheat expressing an antifungal defensin offers a promising approach to complement the endogenous resistance genes within the wheat germplasm for durable resistance to Pt...
August 31, 2016: Transgenic Research
Karin Fister, Iztok Fister, Jana Murovec, Borut Bohanec
Plant breeders' rights are undergoing dramatic changes due to changes in patent rights in terms of plant variety rights protection. Although differences in the interpretation of »breeder's exemption«, termed research exemption in the 1991 UPOV, did exist in the past in some countries, allowing breeders to use protected varieties as parents in the creation of new varieties of plants, current developments brought about by patenting conventionally bred varieties with the European Patent Office (such as EP2140023B1) have opened new challenges...
August 27, 2016: Transgenic Research
Chao Bai, Judit Berman, Gemma Farre, Teresa Capell, Gerhard Sandmann, Paul Christou, Changfu Zhu
Astaxanthin is a high-value ketocarotenoid rarely found in plants. It is derived from β-carotene by the 3-hydroxylation and 4-ketolation of both ionone end groups, in reactions catalyzed by β-carotene hydroxylase and β-carotene ketolase, respectively. We investigated the feasibility of introducing an extended carotenoid biosynthesis pathway into rice endosperm to achieve the production of astaxanthin. This allowed us to identify potential metabolic bottlenecks that have thus far prevented the accumulation of this valuable compound in storage tissues such as cereal grains...
August 27, 2016: Transgenic Research
I M Gerasymenko, Y V Sheludko, A A Klebanovych, V A Rudas, A M Shakhovsky, T M Klein, N V Kuchuk
The development of tools which ensure the desired level of transgene expression in plastids is a prerequisite for the effective utilization of these plant organelles for the deployment of bioactive proteins. High-level accumulation of target proteins is considered as a positive feature of transplastomic plants, but excessive accumulation of foreign proteins may have deleterious effects on host plants. On the other hand, expression at low levels can result in ineffective phenotypes. We compared the effectiveness of different 5'-regulatory sequences in driving the expression of a reporter gene, β-glucuronidase (uidA), in tobacco chloroplasts...
August 26, 2016: Transgenic Research
Dae-Jin Kwon, Dong-Hwan Kim, In-Sul Hwang, Dong-Ern Kim, Hyung-Joo Kim, Jang-Seong Kim, Kichoon Lee, Gi-Sun Im, Jeong-Woong Lee, Seongsoo Hwang
Recent progress in genetic manipulation of pigs designated for xenotransplantation ha6s shown considerable promise on xenograft survival in primates. However, genetic modification of multiple genes in donor pigs by knock-out and knock-in technologies, aiming to enhance immunological tolerance against transplanted organs in the recipients, has not been evaluated for health issues of donor pigs. We produced transgenic Massachusetts General Hospital piglets by knocking-out the α-1,3-galactosyltransferase (GT) gene and by simultaneously knocking-in an expression cassette containing five different human genes including, DAF, CD39, TFPI, C1 inhibitor (C1-INH), and TNFAIP3 (A20) [GT(-(DAF/CD39/TFPI/C1-INH/TNFAIP3)/+)] that are connected by 2A peptide cleavage sequences to release individual proteins from a single translational product...
August 23, 2016: Transgenic Research
Jeanette Orbegozo, Dennis Solorzano, Wilmer J Cuellar, Ida Bartolini, Maria Lupe Roman, Marc Ghislain, Jan Kreuze
An inverted repeat construct corresponding to a segment of the potato leaf roll virus coat protein gene was created under control of a constitutive promoter and transferred into a transformation vector with a heat inducible Cre-loxP system to excise the nptII antibiotic resistance marker gene. Fifty-eight transgenic events were evaluated for resistance to PLRV by greenhouse inoculations, which lead to the identification of 7 highly resistant events, of which 4 were extremely resistant. This resistance was also highly effective against accumulation in subsequent tuber generations from inoculated plants, which has not been reported before...
August 20, 2016: Transgenic Research
Jürgen Breitenbach, Marilise Nogueira, Gemma Farré, Changfu Zhu, Teresa Capell, Paul Christou, Gunther Fleck, Ulfert Focken, Paul D Fraser, Gerhard Sandmann
Astaxanthin from a transgenic maize line was evaluated as feed supplement source conferring effective pigmentation of rainbow trout flesh. An extraction procedure using ethanol together with the addition of vegetal oil was established. This resulted in an oily astaxanthin preparation which was not sufficiently concentrated for direct application to the feed. Therefore, a concentration process involving multiple phase partitioning steps was implemented to remove 90 % of the oil. The resulting astaxanthin raw material contained non-esterified astaxanthin with 12 % 4-keto zeaxanthin and 2 % zeaxanthin as additional carotenoids...
August 12, 2016: Transgenic Research
Mieke Baan, Kathleen J Krentz, Danielle A Fontaine, Dawn Belt Davis
The Black and Tan Brachyury (BTBR) mouse strain is a valuable model for the study of long-term complications from obesity-induced type 2 diabetes mellitus and autism spectrum disorder. Due to technical difficulties with assisted reproduction, genetically modified animals on this background have previously been generated through extensive backcrossing, which is expensive and time-consuming. We successfully generated two separate transgenic mouse lines after direct zygote microinjection into this background strain...
August 11, 2016: Transgenic Research
Oliver G Rössler, Andrea Lesch, Gerald Thiel
The binary tetracycline-based expression system in transgenic mice relies on the expression of the tetracycline transactivator (tTA or rtTA) in a particular cell type together with a transcription unit encoding the gene of interest under a tetracycline or doxycycline-responsive promoter. Transgenic mice containing this transcription unit are produced via pronucleus injection. As the chromosomal integration site of the injected DNA influences transgene expression, several founder lines have to be crossed with (r)tTA-expressing mice to find a line showing low background and high transgene expression following doxycycline stimulation...
August 9, 2016: Transgenic Research
M P Beker, P Boari, M Burachik, V Cuadrado, M Junco, S Lede, M A Lema, D Lewi, A Maggi, I Meoniz, G Noé, C Roca, C Robredo, C Rubinstein, C Vicien, A Whelan
Experience gained in the risk assessment (RA) of genetically engineered (GE) crops since their first experimental introductions in the early nineties, has increased the level of familiarity with these breeding methodologies and has motivated several agencies and expert groups worldwide to revisit the scientific criteria underlying the RA process. Along these lines, the need to engage in a scientific discussion for the case of GE crops transformed with similar constructs was recently identified in Argentina...
October 2016: Transgenic Research
Hengtao Ge, Chenchen Cui, Jun Liu, Yan Luo, Fusheng Quan, Yaping Jin, Yong Zhang
With the technological development of several engineered endonucleases (EENs), such as zinc-finger nucleases, transcription activator-like effector nucleases (TALENs) and CRISPR/Cas9, gene targeting by homologous recombination has been efficiently improved to generate site-specifically genetically modified livestock. However, few studies have been done to investigate the health and fertility of these animals. The purpose of the present study is to investigate if gene targeting events and a recloning procedure would affect the production traits of EEN-mediated gene targeted bucks...
October 2016: Transgenic Research
Mark Tizard, Eric Hallerman, Scott Fahrenkrug, Martina Newell-McGloughlin, John Gibson, Frans de Loos, Stefan Wagner, Götz Laible, Jae Yong Han, Michael D'Occhio, Lisa Kelly, John Lowenthal, Kari Gobius, Primal Silva, Caitlin Cooper, Tim Doran
The ability to generate transgenic animals has existed for over 30 years, and from those early days many predicted that the technology would have beneficial applications in agriculture. Numerous transgenic agricultural animals now exist, however to date only one product from a transgenic animal has been approved for the food chain, due in part to cumbersome regulations. Recently, new techniques such as precision breeding have emerged, which enables the introduction of desired traits without the use of transgenes...
October 2016: Transgenic Research
Natalie J Dorà, Aaron J F Crookshanks, Karen K Y Leung, T Ian Simpson, John O Mason, David J Price, John D West
Analysis of abnormal phenotypes produced by different types of mutations has been crucial for our understanding of gene function. Some floxed alleles that retain a neomycin-resistance selection cassette (neo cassette) are not equivalent to wild-type alleles and provide useful experimental resources. Pax6 is an important developmental gene and the aim of this study was to determine whether the floxed Pax6 (tm1Ued) (Pax6 (fl) ) allele, which has a retained neo cassette, produced any abnormal eye phenotypes that would imply that it differs from the wild-type allele...
October 2016: Transgenic Research
Kaile Sun, Anne-Marie A Wolters, Jack H Vossen, Maarten E Rouwet, Annelies E H M Loonen, Evert Jacobsen, Richard G F Visser, Yuling Bai
Phytophthora infestans, the causal agent of late blight, is a major threat to commercial potato production worldwide. Significant costs are required for crop protection to secure yield. Many dominant genes for resistance (R-genes) to potato late blight have been identified, and some of these R-genes have been applied in potato breeding. However, the P. infestans population rapidly accumulates new virulent strains that render R-genes ineffective. Here we introduce a new class of resistance which is based on the loss-of-function of a susceptibility gene (S-gene) encoding a product exploited by pathogens during infection and colonization...
October 2016: Transgenic Research
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