scholarly journals Embryo Culture, In Vitro Propagation, and Molecular Identification for Advanced Olive Breeding Programs

Horticulturae ◽  
2021 ◽  
Vol 8 (1) ◽  
pp. 36
Author(s):  
Vito Montilon ◽  
Leonardo Susca ◽  
Oriana Potere ◽  
Vincenzo Roseti ◽  
Antonia Campanale ◽  
...  
Keyword(s):  
Seed Germination ◽  
Embryo Culture ◽  
Large Scale ◽  
Olive Tree ◽  
Tree Breeding ◽  
Scale Production ◽  
Breeding Programs ◽  
Time Required ◽  
Olive Breeding

The high biodiversity of the olive tree is an important opportunity to develop sustainable plans to control Xylella fastidiosa (Xf) through breeding programs. Olive tree breeding activities have been limited due to various features of this species including the long time required for seed germination caused by the inhibition effect of the woody endocarp, the seed integument, and the endosperm. Starting from F1 seeds by cross-breeding, the embryo culture was compared with traditional seed germination, evaluating the effectiveness of in vitro multiplication of the plantlets for large-scale production. The isolated embryos were established on a new medium based on Rugini ‘84 macroelements, Murashige & Skoog ‘62 microelements, with Nitsch J. P. & Nitsch C. ‘69 vitamine and subcultured on Leva MSM modified. The results obtained confirmed that in vitro culture of olive embryos is a valid tool for increasing the percentage and speed of germination, helping to reduce the time of the olive breeding programs, offering the possibility to effectively propagate plantlets for further experiments.

scholarly journals Lentiviral Vectors for Delivery of Gene-Editing Systems Based on CRISPR/Cas: Current State and Perspectives

Viruses ◽  
2021 ◽  
Vol 13 (7) ◽  
pp. 1288
Author(s):  
Wendy Dong ◽  
Boris Kantor
Keyword(s):  
Large Scale ◽  
Lentiviral Vector ◽  
Clinical Applications ◽  
Scale Production ◽  
Base Editing ◽  
Epigenome Editing ◽  
Vector Systems ◽  
Dividing Cells

CRISPR/Cas technology has revolutionized the fields of the genome- and epigenome-editing by supplying unparalleled control over genomic sequences and expression. Lentiviral vector (LV) systems are one of the main delivery vehicles for the CRISPR/Cas systems due to (i) its ability to carry bulky and complex transgenes and (ii) sustain robust and long-term expression in a broad range of dividing and non-dividing cells in vitro and in vivo. It is thus reasonable that substantial effort has been allocated towards the development of the improved and optimized LV systems for effective and accurate gene-to-cell transfer of CRISPR/Cas tools. The main effort on that end has been put towards the improvement and optimization of the vector’s expression, development of integrase-deficient lentiviral vector (IDLV), aiming to minimize the risk of oncogenicity, toxicity, and pathogenicity, and enhancing manufacturing protocols for clinical applications required large-scale production. In this review, we will devote attention to (i) the basic biology of lentiviruses, and (ii) recent advances in the development of safer and more efficient CRISPR/Cas vector systems towards their use in preclinical and clinical applications. In addition, we will discuss in detail the recent progress in the repurposing of CRISPR/Cas systems related to base-editing and prime-editing applications.

Abstract 262: Derivation of Duchenne Muscular Dystrophy Disease Specific Cardiomyocytes From Patient Urine

2013 ◽  
Vol 113 (suppl_1) ◽  
Author(s):  
Xuan Guan ◽  
David L Mack ◽  
Claudia M Moreno ◽  
Fernando Santana ◽  
Charles E Murry ◽  
...  
Keyword(s):  
Stem Cells ◽  
Human Urine ◽  
Large Scale ◽  
Lentiviral Vector ◽  
Cellular Reprogramming ◽  
Urine Samples ◽  
Pcr Analysis ◽  
Scale Production ◽  
Mechanism Study

Introduction: Human somatic cells can be reprogrammed into primitive stem cells, termed induced pluripotent stem cells (iPSCs). These iPSCs can be extensively expanded in vitro and differentiated into multiple functional cell types, enabling faithful preservation of individual’s genotype and large scale production of disease targeted cellular components. These unique cellular reagents thus hold tremendous potential in disease mechanism study, drugs screening and cell replacement therapy. Due to the genetic mutation of the protein dystrophin, many DMD patients develop fatal cardiomyopathy with no effective treatment. The underlying pathogenesis has not been fully elucidated. Hypothesis: We tested the hypothesis that iPSCs could be generated from DMD patients’ urine samples and differentiated into cardiomyocytes, recapitulating the dystrophic phenotype. Methods: iPSCs generation was achieved by introducing a lentiviral vector expressing Oct4, Sox2, c-Myc and Klf4 into cells derived from patient’s (n=1) and healthy volunteers’ (n=3) urine. Cardiomyocytes were derived by sequentially treating iPSCs with GSK3 inhibitor CHIR99021 and Wnt inhibitor IWP4. Differentiated cardiomyocytes were subjected to calcium imaging, electrophysiology recording, Polymerase Chain Reaction (PCR) analysis, and immunostaining. Results: iPSCs were efficiently generated from human urine samples and further forced to differentiate into contracting cardiomyocytes. PCR analysis and immunostaining confirmed the expression of a panel of cardiac markers. Both normal and patient iPSC derived cardiomyocytes exhibited spontaneous and field stimulated calcium transients (up to 2Hz), as well as action potentials with ventricular-like and nodal-like characteristics. Anti-dystrophin antibodies stained normal iPSC-derived cardiomyocyte membranes but did not react against DMD iPSC-derived cardiomyocytes. Conclusions: Cardiomyocytes can be efficiently generated from human urine, through the cellular reprogramming technology. DMD cardiomyocytes retained the patient’s genetic information and manifested a dystrophin-null phenotype. Functional assessments are underway to determine differences that may exist between genotypes.

scholarly journals In Vitro Seed Germination and Embryo Culture in Nothapodytes foetida (Wight) Sleumer

2015 ◽  
Vol 48 ◽  
pp. 23-31 ◽  
Author(s):  
S. Kaveri ◽  
Rao Srinath
Keyword(s):  
Seed Germination ◽  
Embryo Culture ◽  
Filter Paper ◽  
Agar Medium ◽  
Ms Medium ◽  
Mature Embryos ◽  
Nothapodytes Foetida ◽  
Half Strength ◽  
Paper Bridge

In vitro seed germination and embryo culture have been achieved in Nothapodytes foetida, this plant is known for its rich source of anticancer drug i. e., Camptothecin. In present study both normal and decoated seeds were subjected to different treatments viz., H2O, GA3, H2O2, H2SO4, chlorine water and mechanical scarification, further these were germinated on water agar medium (WA), filter paper bridge (FB), half strength MS (HMS) and full strength MS (FMS) medium. The highest percentage (69%) of germination was achieved from decoated seeds treated with 10mg/L GA3 and germinated on Filter Paper Bridge. And for embryo culture mature embryos were inoculated on MS medium containing various combination and concentrations of cytokinins (BAP, Kn and TDZ) and auxin (IAA and NAA) for rapid conversion into a plantlet. Among the different combinations of growth regulators; highest frequency (100%) of plantlet conversion was obtained on MS medium containing Kn (1.0mg/L) and NAA (0.2mg/L).

Optimizing in vitro large scale production of giant reed (Arundo donax L.) by liquid medium culture

2014 ◽  
Vol 69 ◽  
pp. 21-27 ◽  
Author(s):  
Valeria Cavallaro ◽  
Cristina Patanè ◽  
Salvatore L. Cosentino ◽  
Isabella Di Silvestro ◽  
Venera Copani
Keyword(s):  
Liquid Medium ◽  
Large Scale ◽  
Arundo Donax ◽  
Giant Reed ◽  
Scale Production ◽  
Large Scale Production ◽  
Medium Culture ◽  
Arundo Donax L

Scalable Microfluidic Platform for Flexible Configuration of and Experiments with Microtissue Multiorgan Models

2018 ◽  
Vol 24 (1) ◽  
pp. 79-95 ◽  
Author(s):  
Christian Lohasz ◽  
Nassim Rousset ◽  
Kasper Renggli ◽  
Andreas Hierlemann ◽  
Olivier Frey
Keyword(s):  
Large Scale ◽  
Plastic Material ◽  
Microtiter Plate ◽  
Scale Production ◽  
Experimental Conditions ◽  
Precise Control ◽  
Open Design ◽  
Gravity Driven ◽  
Gravity Driven Flow

Microphysiological systems hold the promise to increase the predictive and translational power of in vitro substance testing owing to their faithful recapitulation of human physiology. However, the implementation of academic developments in industrial settings remains challenging. We present an injection-molded microfluidic microtissue (MT) culture chip that features two channels with 10 MT compartments each and that was designed in compliance with microtiter plate standard formats. Polystyrene as a chip material enables reliable, large-scale production and precise control over experimental conditions due to low adsorption or absorption of small, hydrophobic molecules at or into the plastic material in comparison with predecessor chips made of polydimethylsiloxane. The chip is operated by tilting, which actuates gravity-driven flow between reservoirs at both ends of every channel, so that the system does not require external tubing or pumps. The flow rate can be modulated by adjusting the tilting angle on demand. The top-open design of the MT compartment enables efficient MT loading using standard or advanced pipetting equipment, ensures oxygen availability in the chip, and allows for high-resolution imaging. Every channel can be loaded with up to 10 identical or different MTs, as demonstrated by culturing liver and tumor MTs in the same medium channel on the chip.

Variations in Pathogenicity of Three Different Forms of Rhizoctonia Inoculum and Assessment of Cultivar Resistance of Sugar Beet

2021 ◽  
Vol 3 (1) ◽  
Author(s):  
Haque ME ◽  
◽  
Parvin MS ◽  
Keyword(s):  
Seed Germination ◽  
Sugar Beet ◽  
Root Rot ◽  
Large Scale ◽  
Culture Media ◽  
Damping Off ◽  
Require Time ◽  
Time Space ◽  
The Usa

Rhizoctonia solani causes pre-emergence and post-emergence damping-off, as well as crown and root rot of sugar beet (Beta vulgaris L.), which significantly affects the yield returns in the USA and Europe. The pathogen overwinters as sclerotia or melanized mycelium. Traditionally, the resistance of cultivars to R. solani is evaluated by scoring disease reactions at the crowns and roots of older seedlings, thus resistance is not evaluated during seed germination. Moreover, earlier studies evaluated cultivars resistance to R. solani using colonized whole barley or wheat grains which, unlike sclerotia, are artificial inocula of the pathogen that require time, space and technical know-how to produce. Moreover, colonized grains are prone to contamination with other pathogens, consumed by rodents/birds while applied in the field, and are often uneconomic. Considering those limitations, a study was undertaken (1) to develop in vitro methods to generate large-scale sclerotia, (2) to compare pathogenic potentials of sclerotia, mycelia, and colonized barley grains for optimization of dampingoff assays, and (3) to evaluate Rhizoctonia resistance of selected commercial cultivars during the seed germination phase. Comparing six different culture media, we found that R. solani had the highest radial growth (8.9 ± 0.04, cm³) at 8-days and the maximum number of sclerotia produced (203 ± 4.6) at 28-days in CV8 medium. We demonstrated significant differences in pathogenicity of the three different forms of R. solani inocula and susceptibility of cultivars to preand post-emergence damping-off. The highest pre-emergence damping-off and root rot were observed with sclerotia, and the highest post-emergence dampingoff was recorded with both sclerotial and colonized barley inocula. In addition, varietal differences in susceptibility to pre- and post-emergence damping-off were noted. The highest pre-emergence damping-off was recorded on cv Crystal 101RR and lowest in Maribo MA 504. The highest post-emergence damping-off was recorded on BTS 8500 and the lowest in Crystal 467. The maximum mean root rot was observed in BTS 8500, BTS 8606, and Crystal 101R. Our studies demonstrated that sclerotia serve as efficient natural inocula, reemphasized that host-pathogen interactions differ at the early vs. late stages of sugar beet growth, and highlighted the need to reevaluate commercial sugar beet cultivars for resistance at the seed germination stage.

Large-scale production of polyoma middle T antigen by using genetically engineered tumors

1985 ◽  
Vol 5 (7) ◽  
pp. 1795-1799
Author(s):  
D R Kaplan ◽  
B Bockus ◽  
T M Roberts ◽  
J Bolen ◽  
M Israel ◽  
...  
Keyword(s):  
Nude Mice ◽  
Large Scale ◽  
T Antigen ◽  
Genetically Engineered ◽  
Scale Production ◽  
Large Scale Production ◽  
Metallothionein Promoter ◽  
Polyoma Middle T Antigen ◽  
Nih 3T3

A recombinant plasmid containing a metallothionein promoter-polyoma middle T cDNA fusion was constructed and used to transfect NIH 3T3 cells. Transformed cells expressing middle T were injected into nude mice. Within 3 weeks, each mouse produced tumors containing middle T equivalent to that in 250 to 1,000 100-mm dishes of polyomavirus-infected cells. This middle T, partially purified by immunoaffinity chromatography, retained activity as measured by its ability to be phosphorylated in vitro. The combined approach of fusing strong promoters to genes of interest and utilizing nude mice to grow large quantities of cells expressing the gene provides a quick, inexpensive alternative to other expression systems.

scholarly journals Production of Phytotoxic Metabolite Using Biphasic Fermentation System from Strain C1136 of Lasiodiplodia pseudotheobromae, a Potential Bioherbicidal Agent

2017 ◽  
Vol 9 (3) ◽  
pp. 371-377
Author(s):  
Charles Oluwaseun ADETUNJI ◽  
Julius Kola OLOKE ◽  
Gandham PRASAD ◽  
Moses ABALAKA ◽  
Emenike Onyebum IROKANULO
Keyword(s):  
Large Scale ◽  
Wild Strain ◽  
Fermentation Medium ◽  
Scale Production ◽  
Conventional Farming ◽  
Large Scale Production ◽  
Phytotoxic Metabolite ◽  
Biphasic Fermentation

Formulation of effective and environmental friendly bioherbicides depends on the type of fermentation medium used for the production of phytotoxic metabolites. The effect of biomass, colony forming unit and the phytotoxic metabolite produced from the biphasic fermentation was carried out, while the phytotoxic metabolite was  tested in vivo and in-vitro on Echinochola crus-galli and dicotyledonous Chromolaena odorata. The mutant strain of Lasiodiplodia pseudotheobromae C1136 (Lp90) produced the highest amount of conidia and the largest necrotic area on the two tested weeds when compared to its wild strain in the different biphasic media combinations. The study revealed that the biphasic system containing PDB + rice produced the highest bioherbicidal activities. Therefore, the phytotoxic metabolites from strain C1136 are suggested for large scale production of bioherbicides for the management of weeds in conventional farming to improve yield and enhance food security.

scholarly journals Expansion of Human Pluripotent Stem Cell-derived Early Cardiovascular Progenitor Cells by a Cocktail of Signaling Factors

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Sadaf Vahdat ◽  
Sara Pahlavan ◽  
Elena Mahmoudi ◽  
Maryam Barekat ◽  
Hassan Ansari ◽  
...  
Keyword(s):  
Progenitor Cells ◽  
Large Scale ◽  
Culture Medium ◽  
Gene Expression Pattern ◽  
Extended Period ◽  
Scale Production ◽  
Chemical Screening ◽  
Wide Range ◽  
Cell Sources

Abstract Cardiovascular progenitor cells (CPCs) derived from human pluripotent stem cells (hPSCs) are proposed to be invaluable cell sources for experimental and clinical studies. This wide range of applications necessitates large-scale production of CPCs in an in vitro culture system, which enables both expansion and maintenance of these cells. In this study, we aimed to develop a defined and efficient culture medium that uses signaling factors for large-scale expansion of early CPCs, called cardiogenic mesodermal cells (CMCs), which were derived from hPSCs. Chemical screening resulted in a medium that contained a reproducible combination of three factors (A83-01, bFGF, and CHIR99021) that generated 1014 CMCs after 10 passages without the propensity for tumorigenicity. Expanded CMCs retained their gene expression pattern, chromosomal stability, and differentiation tendency through several passages and showed both the safety and possible cardio-protective potentials when transplanted into the infarcted rat myocardium. These CMCs were efficiently cryopreserved for an extended period of time. This culture medium could be used for both adherent and suspension culture conditions, for which the latter is required for large-scale CMC production. Taken together, hPSC-derived CMCs exhibited self-renewal capacity in our simple, reproducible, and defined medium. These cells might ultimately be potential, promising cell sources for cardiovascular studies.

scholarly journals Regeneration Technique of Bamboo Species through Nodal Segments: A Review

2020 ◽  
Vol 8 (1) ◽  
pp. 54-68
Author(s):  
Meena Maiya Suwal ◽  
Janardan Lamichhane ◽  
Dhurva Prasad Gauchan
Keyword(s):  
Large Scale ◽  
Physiological Stress ◽  
Perennial Grass ◽  
Shoot Multiplication ◽  
Nodal Segments ◽  
Individual Species ◽  
Small Scale ◽  
Scale Production ◽  
Bamboo Species

Micropropagation is an alternative technique to propagate at large scale plants to meet global plant demand. Various researchers have worked on the micropropagation technique to regenerate bamboo species by using nodal segments from years. Contamination, browning, necrosis, and acclimatization with physiological stress are the extreme problems of the micropropagation technique. But, many numbers of papers have been published on micropropagation of the bamboo species through nodal segments as explants. The proliferation of the bamboo shoots is dependent on the season of collection, size of explants, the position of explants, diversity of plants, concentration and combination of plant growth regulators, most adequate culture medium, environmental condition of the equipment, handling, and individual species. Bamboo is a monocarpic fast-growing, tall perennial grass and having the high potential to generate economic and social benefits. It helps to maintain land patterns and control soil erosion.  The long life cycle of the bamboo produces a huge amount of seeds but unfortunately, mostly, they are non-viable. So, bamboos are propagated from vegetative by cutting and air layering. However, these methods are only for a small scale and they also tend to destroy large mother plant stocks and difficult to be transported. So, the in vitro propagation technique is useful to obtain large progenies from desired genotypes. Mostly, BAP and TDZ growth hormones are widely used for shoot multiplication and IBA, NAA and IAA are used for root initiation as per developed protocols in tissue culture for large scale production. This review intends to explore an overview of the recent literature reports to summarize the importance of micropropagation by using nodal segments of bamboo species and factors influencing it.

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