Major Techniques of Biotechnology

1998 ◽  
Vol 11 (1) ◽  
pp. 28-37
Author(s):  
Joseph A. Tami
Keyword(s):  
Messenger Rna ◽  
Recombinant Dna ◽  
Inflammatory Diseases ◽  
Diagnostic Testing ◽  
Yeast Cells ◽  
Recombinant Dna Technology ◽  
Potential Applications ◽  
Foreign Dna ◽  
Immuno Globulins ◽  
And Function

Since the discovery of the structure and function of DNA over 40 years ago, the established knowledge of molecular biology has increased dramatically, and many new tools have been discovered and utilized by scientists to develop new therapeutic agents. Important tools that are used in recombinant DNA technology include restriction endonucleases (cleave DNA), DNA ligase (link DNA molecules together), and cloning vectors (place foreign DNA into an organism such as bacterial or yeast cells in order to mass produce the protein encoded by that foreign DNA). The development of hybridoma technology provided a method to produce virtually unlimited quantities of pure antibody with a single specificity. These immuno-globulins are known as monoclonal antibodies, and have provided both therapeutic and diagnostic agents. Antisense molecules are oligonucleotides which bind to the messenger RNA (mRNA) of a target gene and selectively inhibit the production of specific proteins. Potential applications for these molecules include cancer and viral and inflammatory diseases. The more recent development of the polymerase chain reaction (PCR) has provided a tool that has revolutionized diagnostic testing in diverse areas such as infectious diseases, genetic abnormalities, and cancer.

scholarly journals Recombinant DNA technology and click chemistry: a powerful combination for generating a hybrid elastin-like-statherin hydrogel to control calcium phosphate mineralization

2017 ◽  
Vol 8 ◽  
pp. 772-783 ◽  
Author(s):  
Mohamed Hamed Misbah ◽  
Mercedes Santos ◽  
Luis Quintanilla ◽  
Christina Günter ◽  
Matilde Alonso ◽  
...  
Keyword(s):  
Calcium Phosphate ◽  
Click Chemistry ◽  
Recombinant Dna ◽  
Fibrillar Structure ◽  
Cross Link ◽  
Recombinant Dna Technology ◽  
Peptide Epitope ◽  
Phosphate Mineralization ◽  
Potential Applications ◽  
Dna Technology

Understanding the mechanisms responsible for generating different phases and morphologies of calcium phosphate by elastin-like recombinamers is supreme for bioengineering of advanced multifunctional materials. The generation of such multifunctional hybrid materials depends on the properties of their counterparts and the way in which they are assembled. The success of this assembly depends on the different approaches used, such as recombinant DNA technology and click chemistry. In the present work, an elastin-like recombinamer bearing lysine amino acids distributed along the recombinamer chain has been cross-linked via Huisgen [2 + 3] cycloaddition. The recombinamer contains the SNA15 peptide domains inspired by salivary statherin, a peptide epitope known to specifically bind to and nucleate calcium phosphate. The benefit of using click chemistry is that the hybrid elastin-like-statherin recombinamers cross-link without losing their fibrillar structure. Mineralization of the resulting hybrid elastin-like-statherin recombinamer hydrogels with calcium phosphate is described. Thus, two different hydroxyapatite morphologies (cauliflower- and plate-like) have been formed. Overall, this study shows that crosslinking elastin-like recombinamers leads to interesting matrix materials for the generation of calcium phosphate composites with potential applications as biomaterials.

Gene Cloning

2021 ◽  
pp. 165-218
Keyword(s):  
Recombinant Dna ◽  
Restriction Enzymes ◽  
Protein Product ◽  
Biological Molecules ◽  
Recombinant Dna Technology ◽  
Phage Dna ◽  
Controlling Elements ◽  
Foreign Dna ◽  
Dna Fragment ◽  
Dna Technology

The discovery of two naturally occurring biological molecules, plasmid DNA and restriction enzymes, with remarkable properties have made possible the development of methods to isolate and manipulate specific DNA fragments. Through this technology, a DNA fragment, even an entire gene and its controlling elements, can be isolated and rejoined with a plasmid or phage DNA, and the hybrid DNA molecule can be inserted into a bacterium. The foreign DNA insert can be multiplied inside the bacterial host and induced to express or synthesize the protein product of the foreign DNA. The entire process through which this can be achieved is called recombinant DNA technology or genetic engineering. The recombinant DNA technology has been extended to animal and plant cells. In this chapter, methods for isolation, modification, rejoining and replication of genomic DNA, and production of new or enhanced protein products within a host cell have been described.

Examining the Possible Molecular Origins for Enamel Protein Complexity

1987 ◽  
Vol 1 (2) ◽  
pp. 298-305 ◽  
Author(s):  
M.L. Snead ◽  
E.C. Lau
Keyword(s):  
Messenger Rna ◽  
Recombinant Dna ◽  
Germ Line ◽  
Nucleic Acid Hybridization ◽  
Northern Analysis ◽  
Recombinant Dna Technology ◽  
Dental Tissue ◽  
Free System ◽  
Amelogenin Gene ◽  
Translation Apparatus

Our strategy was to examine each of the three loci capable of contributing to the observed complexity 0 of mouse amelogenin proteins recovered from forming enamel: the genome (gene); the transcription apparatus (messenger RNA); and the translation apparatus (proteins). Our approach was based on recombinant DNA technology and a complementary DNA (cDNA) clone, pMa5-5, specific to the predominant mouse amelogenin protein. An "artificial ameloblast" was engineered based on pMa5-5 and the resulting synthetic products compared to those from authentic ameloblasts. First, the genome probably is not responsible for amelogenin complexity: Southern analysis indicates that the amelogenin gene exists as a single copy in either differentiated dental tissue or germ line tissue. Thus, ectomesenchymal-derived instructive signals for ameloblast differentiation do not lead to re-arrangement or amplification of the amelogenin gene. Next, using nucleic acid hybridization techniques, we examined messenger RNA from mouse ameloblasts. Northern analysis of authentic mRNA from mouse ameloblasts, with either the intact or 3'-end of pMa 5-5 used as the reporter molecule, indicates that only one size class of mRNA was detectable. We conclude that at the sensitivity of this assay there is no evidence for multiple mRNAs. Last, "artificial ameloblasts" were engineered so that the translation apparatus could be examined as a source of amelogenin complexity. Capped, artificial mRNAs were constructed to the pMa 5-5 template and used to program the synthesis of amelogenin polypeptides by translation in a cell-free system. When the resulting total translation products were immunoprecipitated with the rabbit anti-mouse amelogenin antibody, we observed multiple polypeptides, suggesting that the utilization of alternative start sites may also contribute to the observed complexity of amelogenin proteins, at least for artificial mRNAs translated in vitro.

scholarly journals A Simple Methodology for Conversion of Mouse Monoclonal Antibody to Human-Mouse Chimeric Form

2013 ◽  
Vol 2013 ◽  
pp. 1-6 ◽  
Author(s):  
Vinh T. Dang ◽  
Kedar D. Mandakhalikar ◽  
Oi-Wing Ng ◽  
Yee-Joo Tan
Keyword(s):  
Monoclonal Antibody ◽  
Recombinant Dna ◽  
Molecular Techniques ◽  
Human Beings ◽  
Recombinant Dna Technology ◽  
Passive Immunotherapy ◽  
Inflammatory Conditions ◽  
Mechanism Of Inhibition ◽  
Set Up ◽  
And Function

Passive immunotherapy has mainly been used as a therapy against cancer and inflammatory conditions. Recent studies have shown that monoclonal antibody-(mAb-) based passive immunotherapy is a promising approach to combat virus infection. Specific mouse mAbs can be routinely generated in large amounts with the use of hybridoma technology but these cannot be used for therapy in human beings due to their immunogenicity. Therefore, the development of chimeric and humanized mAbs is important for therapeutic purpose. This is facilitated by a variety of molecular techniques like recombinant DNA technology and the better understanding of the structure and function of antibody. The human-mouse chimeric forms allow detailed analysis of the mechanism of inhibition and the potential for therapeutic applications. Here, a step-by-step description of the conversion process will be described. The commercial availability of the reagents required in each step means that this experimentation can be easily set up in research laboratories.

scholarly journals Genetic Engineering Technique in Virus-Like Particle Vaccine Construction

2021 ◽  
Vol 16 (4) ◽  
pp. 203
Author(s):  
Debie Rizqoh
Keyword(s):  
Immune Responses ◽  
Mammalian Cells ◽  
Recombinant Dna ◽  
Cytotoxic T Cells ◽  
Yeast Cells ◽  
Expression Systems ◽  
Recombinant Dna Technology ◽  
Recombinant Gene Expression ◽  
Humoral Immune ◽  
Humoral Immune Responses

Vaccine becomes a very effective strategy to deal with various infectious diseases even to the point of eradication as in the smalpox virus. At present many conventional vaccines such as inactivated and live-attenuated vaccines. However, these vaccine methods have side effects on the population. Viral-like particle (VLP) is an alternative vaccine based on recombinant DNA technology that is safe with the same immunogenicity as conventional viruses. This vaccine has been shown to induce humoral immune responses mediated by antibodies and cellular immune responses mediated by cytotoxic T cells. With these advantages, currently various types of vaccines have only been developed on a VLP basis. VLP can be produced from a variety of recombinant gene expression systems including bacterial cell expression systems, yeast cells, insect cells, mammalian cells, plant cells, and cell-free systems.

Enzymatic Synthesis of 15N-L-aspartic Acid Using Recombinant Aspartase from Escherichia Coli K12

2008 ◽  
Vol 59 (11) ◽  
Author(s):  
Iulia Lupan ◽  
Sergiu Chira ◽  
Maria Chiriac ◽  
Nicolae Palibroda ◽  
Octavian Popescu
Keyword(s):  
Amino Acids ◽  
Aspartic Acid ◽  
Enzymatic Synthesis ◽  
Large Scale ◽  
Recombinant Dna ◽  
Industrial Enzymes ◽  
Recombinant Dna Technology ◽  
Bacterial Fermentation ◽  
Natural Protein ◽  
Novel Method

Amino acids are obtained by bacterial fermentation, extraction from natural protein or enzymatic synthesis from specific substrates. With the introduction of recombinant DNA technology, it has become possible to apply more rational approaches to enzymatic synthesis of amino acids. Aspartase (L-aspartate ammonia-lyase) catalyzes the reversible deamination of L-aspartic acid to yield fumaric acid and ammonia. It is one of the most important industrial enzymes used to produce L-aspartic acid on a large scale. Here we described a novel method for [15N] L-aspartic synthesis from fumarate and ammonia (15NH4Cl) using a recombinant aspartase.

Establishing a model to demonstrate physical and mathematical properties of chromatin fibres in fission yeast cells - Research in the Molecular Biophysics Lab at Hiroshima University

Impact ◽  
2018 ◽  
Vol 2018 (3) ◽  
pp. 89-91
Author(s):  
Shin-ichi Tate
Keyword(s):  
Molecular Biology ◽  
Yeast Cells ◽  
Molecular Architecture ◽  
Ministry Of Education ◽  
Cellular Functions ◽  
Sports Science ◽  
Cellular Events ◽  
And Function ◽  
Education Culture ◽  
Open The Doors

The field of molecular biology has provided great insights into the structure and function of key molecules. Thanks to this area of research, we can now grasp the biological details of DNA and have characterised an enormous number of molecules in massive data bases. These 'biological periodic tables' have allowed scientists to connect molecules to particular cellular events, furthering scientific understanding of biological processes. However, molecular biology has yet to answer questions regarding 'higher-order' molecular architecture, such as that of chromatin. Chromatin is the molecular material that serves as the building block for chromosomes, the structures that carry an organism's genetic information inside of the cell's nucleus. Understanding the physical properties of chromatin is crucial in developing a more thorough picture of how chromatin's structure relate to its key cellular functions. Moreover, by establishing a physical model of chromatin, scientists will be able to open the doors into the true inner workings of the cell nucleus. Professor Shin-ichi Tate and his team of researchers at Hiroshima University's Research Center for the Mathematics on Chromatin Live Dynamics (RcMcD), are attempting to do just that. Through a five-year grant funded by the Platform for Dynamic Approaches to Living Systems from the Ministry of Education, Culture, Sports, Science and Technology, Tate is aiming to gain a clearer understanding of the structure and dynamics of chromatin.

scholarly journals Protein-Engineered Polymers Functionalized with Antimicrobial Peptides for the Development of Active Surfaces

2021 ◽  
Vol 11 (12) ◽  
pp. 5352
Author(s):  
Ana Margarida Pereira ◽  
Diana Gomes ◽  
André da Costa ◽  
Simoni Campos Dias ◽  
Margarida Casal ◽  
...  
Keyword(s):  
Antimicrobial Activity ◽  
Antimicrobial Peptides ◽  
Recombinant Dna ◽  
Biological Properties ◽  
Resistant Bacteria ◽  
Recombinant Dna Technology ◽  
Free Standing ◽  
Microbial Cell Factories ◽  
Cell Factories ◽  
Antimicrobial Materials

Antibacterial resistance is a major worldwide threat due to the increasing number of infections caused by antibiotic-resistant bacteria with medical devices being a major source of these infections. This suggests the need for new antimicrobial biomaterial designs able to withstand the increasing pressure of antimicrobial resistance. Recombinant protein polymers (rPPs) are an emerging class of nature-inspired biopolymers with unique chemical, physical and biological properties. These polymers can be functionalized with antimicrobial molecules utilizing recombinant DNA technology and then produced in microbial cell factories. In this work, we report the functionalization of rPBPs based on elastin and silk-elastin with different antimicrobial peptides (AMPs). These polymers were produced in Escherichia coli, successfully purified by employing non-chromatographic processes, and used for the production of free-standing films. The antimicrobial activity of the materials was evaluated against Gram-positive and Gram-negative bacteria, and results showed that the polymers demonstrated antimicrobial activity, pointing out the potential of these biopolymers for the development of new advanced antimicrobial materials.

scholarly journals Expression and function of Smad7 in autoimmune and inflammatory diseases

2021 ◽  
Author(s):  
Yiping Hu ◽  
Juan He ◽  
Lianhua He ◽  
Bihua Xu ◽  
Qingwen Wang
Keyword(s):  
Posttranscriptional Regulation ◽  
Molecular Mechanisms ◽  
Transforming Growth Factor ◽  
Inflammatory Diseases ◽  
Kidney Diseases ◽  
Critical Role ◽  
Transforming Growth Factor Β ◽  
Negative Regulator ◽  
Signaling Mechanism ◽  
And Function

AbstractTransforming growth factor-β (TGF-β) plays a critical role in the pathological processes of various diseases. However, the signaling mechanism of TGF-β in the pathological response remains largely unclear. In this review, we discuss advances in research of Smad7, a member of the I-Smads family and a negative regulator of TGF-β signaling, and mainly review the expression and its function in diseases. Smad7 inhibits the activation of the NF-κB and TGF-β signaling pathways and plays a pivotal role in the prevention and treatment of various diseases. Specifically, Smad7 can not only attenuate growth inhibition, fibrosis, apoptosis, inflammation, and inflammatory T cell differentiation, but also promotes epithelial cells migration or disease development. In this review, we aim to summarize the various biological functions of Smad7 in autoimmune diseases, inflammatory diseases, cancers, and kidney diseases, focusing on the molecular mechanisms of the transcriptional and posttranscriptional regulation of Smad7.

scholarly journals Extracellular vesicles as mediators and markers of acute organ injury: current concepts

2021 ◽  
Author(s):  
Birte Weber ◽  
Niklas Franz ◽  
Ingo Marzi ◽  
Dirk Henrich ◽  
Liudmila Leppik
Keyword(s):  
Extracellular Vesicles ◽  
Inflammatory Diseases ◽  
Organ Injury ◽  
Isolation And Characterization ◽  
Communication Processes ◽  
Incidence And Mortality ◽  
New Biomarkers ◽  
And Function

AbstractDue to the continued high incidence and mortality rate worldwide, there is a need to develop new strategies for the quick, precise, and valuable recognition of presenting injury pattern in traumatized and poly-traumatized patients. Extracellular vesicles (EVs) have been shown to facilitate intercellular communication processes between cells in close proximity as well as distant cells in healthy and disease organisms. miRNAs and proteins transferred by EVs play biological roles in maintaining normal organ structure and function under physiological conditions. In pathological conditions, EVs change the miRNAs and protein cargo composition, mediating or suppressing the injury consequences. Therefore, incorporating EVs with their unique protein and miRNAs signature into the list of promising new biomarkers is a logical next step. In this review, we discuss the general characteristics and technical aspects of EVs isolation and characterization. We discuss results of recent in vitro, in vivo, and patients study describing the role of EVs in different inflammatory diseases and traumatic organ injuries. miRNAs and protein signature of EVs found in patients with acute organ injury are also debated.

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