scholarly journals Homologous alignment cloning: a rapid, flexible and highly efficient general molecular cloning method

PeerJ ◽  
2018 ◽  
Vol 6 ◽  
pp. e5146 ◽  
Author(s):  
Lendl Tan ◽  
Emily J. Strong ◽  
Kyra Woods ◽  
Nicholas P. West
Keyword(s):  
Molecular Cloning ◽  
Low Cost ◽  
Plasmid Vector ◽  
High Yield ◽  
Exonuclease Activity ◽  
Reaction Products ◽  
Simultaneous Exposure ◽  
Highly Efficient ◽  
Cloning Method ◽  
Homologous Alignment

Homologous alignment cloning (HAC) is a rapid method of molecular cloning that facilitates low-cost, highly efficient cloning of polymerase chain reaction products into any plasmid vector in approximately 2 min. HAC facilitates insert integration due to a sequence alignment strategy, by way of short, vector-specific homology tails appended to insert during amplification. Simultaneous exposure of single-stranded fragment ends, utilising the 3′→5′ exonuclease activity of T4 DNA polymerase, creates overlapping homologous DNA on each molecule. The exonuclease activity of T4 polymerase is quenched simply by the addition of EDTA and a simple annealing step ensures high yield and high fidelity vector formation. The resultant recombinant plasmids are transformed into standardE. colicloning strains and screened via established methods as necessary. HAC exploits reagents commonly found in molecular research laboratories and achieves efficiencies that exceed conventional cloning methods, including another ligation-independent method we tested. HAC is also suitable for combining multiple fragments in a single reaction, thus extending its flexibility.

scholarly journals Mega primer-mediated molecular cloning strategy for chimaeragenesis and long DNA fragment insertion

2017 ◽  
Vol 37 (2) ◽  
Author(s):  
Hui Zhang ◽  
Chang-Jun Liu ◽  
Hui Jiang ◽  
Lu Zhou ◽  
Wen-Ying Li ◽  
...  
Keyword(s):  
Molecular Cloning ◽  
High Efficiency ◽  
Low Cost ◽  
Plasmid Vector ◽  
Efficient Tool ◽  
Overlap Extension ◽  
Cloning Strategy ◽  
Dna Fragment ◽  
Fragment Insertion ◽  
Dpni Digestion

Molecular cloning methods based on primer and overlap-extension PCR are widely used due to their simplicity, reliability, low cost and high efficiency. In this article, an efficient mega primer-mediated (MP) cloning strategy for chimaeragenesis and long DNA fragment insertion is presented. MP cloning is a seamless, restriction/ligation-independent method that requires only three steps: (i) the first PCR for mega primer generation; (ii) the second PCR for exponential amplification mediated by the mega primers and (iii) DpnI digestion and transformation. Most importantly, for chimaeragenesis, genes can be assembled and constructed into the plasmid vector in a single PCR step. By employing this strategy, we successfully inserted four DNA fragments (approximately 500 bp each) into the same vector simultaneously. In conclusion, the strategy proved to be a simple and efficient tool for seamless cloning.

Visible Light Assisted Hantzsch Reaction: Synthesis of Polycyclic Dihydropyridines

2019 ◽  
Vol 16 (8) ◽  
pp. 676-682
Author(s):  
Ankusab Noorahmadsab Nadaf ◽  
Kalegowda Shivashankar
Keyword(s):  
Reaction Time ◽  
Light Intensity ◽  
Ammonium Acetate ◽  
Low Cost ◽  
Aromatic Aldehydes ◽  
Light Irradiation ◽  
High Yield ◽  
Short Reaction Time ◽  
Green Protocol ◽  
Dihydropyridine Derivatives

The polycyclic dihydropyridine nucleus represents the heterocyclic system of invaluable core motifs with wide applications in chemical, biological and physical properties. Although this kind of compounds have been extensively synthesized by other groups, the synthesis of these compounds under CFL light intensity were not explored. The synthesis of polycyclic dihydropyridine derivatives were achieved through the reaction of 4-hydroxycoumarin, aromatic aldehydes and ammonium acetate under CFL light irradiation conditions. A series of polycyclic dihydropyridine derivatives were prepared under CFL light irradiation conditions with high yield, short reaction time, ambient condition and without the use of catalyst. The results displayed an efficient method for the synthesis of polycyclic dihydropyridine derivatives. Clean profile, short reaction time, low cost and use of CFL light intensity instead of catalyst making it a genuinely green protocol.

Using Low Temperature, Cost-effective and Green Methods to Carbon Nanohorn Synthesis

2019 ◽  
Vol 9 (2) ◽  
pp. 157-160
Author(s):  
Ali Hasani
Keyword(s):  
Laser Ablation ◽  
Melting Point ◽  
Palm Olein ◽  
Arc Discharge ◽  
Low Cost ◽  
The Other ◽  
High Yield ◽  
Reaction Field ◽  
Carbon Nanohorns ◽  
Graphite Rod

Background: Laser ablation method has high-yield and pure SWCNHs. On the other hand, arc discharge methods have low-cost production of SWCNHs. However, these techniques have more desirable features, they need special expertness to use high power laser or high current discharge that either of them produces very high temperature. As for the researches, the temperatures of these techniques are higher than 4727°C to vaporize the graphite. So, to become aware of the advantages of SWCNHs, it is necessary to find a new way to synthesize SWCNHs at a lower temperature. In other words, reaction field can be expandable at a moderate temperature. This paper reports a new way to synthesize SWCNHs at an extremely reduced temperature. Methods: According to this study, the role of N2 is the protection of the copper holder supporting the graphite rod by increasing heat transfer from the holder. After the current of 70 A was supplied to the system, the temperature of graphite rod was raised to 1600°C. It is obvious that this temperature is somehow higher than the melting point of palladium, 1555°C, and much lower than graphite melting point, 3497°C. Results: Based on the results, there are transitional precursors simultaneous with the SWCNHs. This composition can be created by distortion of the primary SWCNTs at the higher temperature. Subsequently, each SWCNTs have a tendency to be broken into individual horns. With increasing the concentration of the free horns, bud-like SWCNHs can be produced. Moreover, there are individual horns almost separated from the mass of single wall carbon nanohorns. This structure is not common in SWCNHs synthesized by the usual method such as arc discharge or laser ablation. Through these regular techniques, SWCNHs are synthesized as cumulative particles with diameters about 30-150 nm. Conclusion: A simple heating is needed for SWCNTs transformation to SWCNHs with the presence of palladium as catalyst. The well-thought-out mechanism for this transformation is that SWCNTs were initially changed to highly curled shape, and after that were formed into small independent horns. The other rout to synthesize SWCNHs is the pyrolysis of palm olein at 950°C with the assistance of zinc nitrate and ferrocene. Palm olein was used as a promising, bio-renewable and inexpensive carbon source for the production of carbon nanohorns.

scholarly journals Generation of High-Yield, Functional Oligodendrocytes from a c-myc Immortalized Neural Cell Line, Endowed with Staminal Properties

2021 ◽  
Vol 22 (3) ◽  
pp. 1124
Author(s):  
Mafalda Giovanna Reccia ◽  
Floriana Volpicelli ◽  
Eirkiur Benedikz ◽  
Åsa Fex Svenningsen ◽  
Luca Colucci-D’Amato
Keyword(s):  
Cell Line ◽  
Low Cost ◽  
Neural Cell ◽  
New Drugs ◽  
High Yield ◽  
Time Saving ◽  
Interacting Protein ◽  
Neuronal Marker ◽  
Differentiation Status

Neural stem cells represent a powerful tool to study molecules involved in pathophysiology of Nervous System and to discover new drugs. Although they can be cultured and expanded in vitro as a primary culture, their use is hampered by their heterogeneity and by the cost and time needed for their preparation. Here we report that mes-c-myc A1 cells (A1), a neural cell line, is endowed with staminal properties. Undifferentiated/proliferating and differentiated/non-proliferating A1 cells are able to generate neurospheres (Ns) in which gene expression parallels the original differentiation status. In fact, Ns derived from undifferentiated A1 cells express higher levels of Nestin, Kruppel-like factor 4 (Klf4) and glial fibrillary protein (GFAP), markers of stemness, while those obtained from differentiated A1 cells show higher levels of the neuronal marker beta III tubulin. Interestingly, Ns differentiation, by Epidermal Growth Factors (EGF) and Fibroblast Growth Factor 2 (bFGF) withdrawal, generates oligodendrocytes at high-yield as shown by the expression of markers, Galactosylceramidase (Gal-C) Neuron-Glial antigen 2 (NG2), Receptor-Interacting Protein (RIP) and Myelin Basic Protein (MBP). Finally, upon co-culture, Ns-A1-derived oligodendrocytes cause a redistribution of contactin-associated protein (Caspr/paranodin) protein on neuronal cells, as primary oligodendrocytes cultures, suggesting that they are able to form compact myelin. Thus, Ns-A1-derived oligodendrocytes may represent a time-saving and low-cost tool to study the pathophysiology of oligodendrocytes and to test new drugs.

MOF-derived M-OOH with rich oxygen defects by in-situ electro-oxidation reconstitution for highly efficient oxygen evolution reaction

2021 ◽  
Author(s):  
Zhikai Shi ◽  
Zebin Yu ◽  
Ronghua Jiang ◽  
Jun Huang ◽  
Yanping Hou ◽  
...  
Keyword(s):  
Oxygen Evolution ◽  
Oxygen Evolution Reaction ◽  
Energy Production ◽  
Low Cost ◽  
Highly Efficient ◽  
Oxygen Defects ◽  
Electro Oxidation

The oxygen evolution reaction (OER) is an important half-reaction in the field of energy production. However, how effectively, simply, and greenly to prepare low-cost OER electrocatalysts remains a problem. Herein,...

High‐Yield and Low‐Cost Solar Water Purification via Hydrogel‐Based Membrane Distillation

2021 ◽  
pp. 2101036
Author(s):  
Hengyi Lu ◽  
Wen Shi ◽  
Fei Zhao ◽  
Wenjing Zhang ◽  
Peixin Zhang ◽  
...  
Keyword(s):  
Water Purification ◽  
Low Cost ◽  
Membrane Distillation ◽  
High Yield ◽  
Solar Water

scholarly journals Design of Low-Cost and High-Strength Titanium Alloys Using Pseudo-Spinodal Mechanism through Diffusion Couple Technology and CALPHAD

Materials ◽  
2021 ◽  
Vol 14 (11) ◽  
pp. 2910
Author(s):  
Chaoyi Ding ◽  
Chun Liu ◽  
Ligang Zhang ◽  
Di Wu ◽  
Libin Liu
Keyword(s):  
Diffusion Couple ◽  
Titanium Alloys ◽  
High Throughput ◽  
Volume Fraction ◽  
Raw Materials ◽  
Low Cost ◽  
High Strength ◽  
High Yield ◽  
Α Phase ◽  
Cr System

The high cost of development and raw materials have been obstacles to the widespread use of titanium alloys. In the present study, the high-throughput experimental method of diffusion couple combined with CALPHAD calculation was used to design and prepare the low-cost and high-strength Ti-Al-Cr system titanium alloy. The results showed that ultra-fine α phase was obtained in Ti-6Al-10.9Cr alloy designed through the pseudo-spinodal mechanism, and it has a high yield strength of 1437 ± 7 MPa. Furthermore, application of the 3D strength model of Ti-6Al-xCr alloy showed that the strength of the alloy depended on the volume fraction and thickness of the α phase. The large number of α/β interfaces produced by ultra-fine α phase greatly improved the strength of the alloy but limited its ductility. Thus, we have demonstrated that the pseudo-spinodal mechanism combined with high-throughput diffusion couple technology and CALPHAD was an efficient method to design low-cost and high-strength titanium alloys.

Embedding metal foam into metal–organic framework monoliths for triggering a highly efficient release of adsorbed atmospheric water by localized eddy current heating

2021 ◽  
Author(s):  
Yingle Tao ◽  
Qiangqiang Li ◽  
Qiannan Wu ◽  
Haiqing Li
Keyword(s):  
Thermal Insulation ◽  
Eddy Current ◽  
Energy Efficient ◽  
Metal Foam ◽  
Low Cost ◽  
Metal Organic Framework ◽  
Atmospheric Water ◽  
Highly Efficient ◽  
Metal Organic ◽  
Eddy Current Heating

Localized eddy current heating delivered by metal foam embedded in a MOF monolith provides a novel, low-cost, and energy efficient way to overcome the thermal insulation nature of MOF monoliths and realize their highly efficient regenerations.

A very simple and high-yield method to synthesize nanolayered Mn oxide

2015 ◽  
Vol 44 (3) ◽  
pp. 1039-1045 ◽  
Author(s):  
Mohammad Mahdi Najafpour ◽  
Emad Amini
Keyword(s):  
Low Cost ◽  
High Yield ◽  
Mn Oxide ◽  
Mn Oxides

Nanolayered Mn oxides have been prepared by a very simple, low-cost and high-yield method using soap, KOH, MnCl2and H2O2.

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