scholarly journals Kloning dan Analisis Bioinformatika Gen MSP1 Plasmodium falciparum Isolat Kota Jayapura

2018 ◽  
Vol 10 (1) ◽  
Author(s):  
Arsyam Mawardi ◽  
Leonardo E. Aisoi ◽  
Paula N. Lefaan
Keyword(s):  
Plasmodium Falciparum ◽  
Heat Shock ◽  
Phylogenetic Tree ◽  
Recombinant Plasmid ◽  
Bioinformatics Analysis ◽  
Competent Cell ◽  
E Coli ◽  
Competent Cells ◽  
Plasmid Isolation ◽  
Msp1 Gene

Cloning gene involves the construction of a recombinant plasmid that inserted in a competent cell. On the other hand, genetic engineering requires bioinformatic analysis to be converted into tabulation and data interpretation. The study, titled "cloning block 2 MSP1 gene of Plasmodium falciparum isolate Jayapura city and bioinformatics analysis" is aimed to improve the technique of cloning the MSP1 gene of P. falciparum, initiated the creation of DH5α competent cells, ligations and transformations, plasmid isolation, confirmation the recombinant plasmid and able to perform bioinformatics analysis and construct phylogenetic tree. This study began with the manufacture of E. coli DH5α competent cells, MSP1 gene ligation in pJET1.2/blunt vector and transformation by using the heat shock transformation method, plasmid isolation of alkali lysis method, then plasmid confirmed by PCR and sequencing method, further sequence analysis and phylogenetic tree construction. The results showed that confirmation of MSP1 gene presence in pJET1.2/blunt with PCR was successful. From a total of 4 positive colonies grown in liquid culture, then isolated plasmid and confirmed with PCR obtained electroferogram bands with a size about 1049 bp indicates the presence of MSP1 gene in plasmid. Based on the results, cloning of MSP1 gene using pJET1.2/blunt cloning vector and competent cell E. coli DH5α has been successfully performed. Bioinformatics analysis of sequencing result and phylogenetic tree were constructed successfully with 2 clusters isolate of malaria patients from Jayapura city. Key words: Bioinformatics, cloning gene, heat shock transformation, MSP1, P. falciparum.

scholarly journals Molecular Diversity of Bacillus thuringiensis and Bioinformatics Analysis of Local Isolate of Auky Island, Padaido District in Biak Numfor Papua as a Control of Anopheles Mosquito Larvae

2021 ◽  
Author(s):  
Daniel Lantang ◽  
Arsyam Mawardi
Keyword(s):  
Molecular Diversity ◽  
Bioinformatics Analysis ◽  
Anopheles Mosquito ◽  
Mosquito Larvae ◽  
Sequencing Analysis ◽  
E Coli ◽  
16S Gene ◽  
Tree Construction ◽  
Pcr Product ◽  
Plasmid Isolation

This research aims to analyze the level of similarity and diversity among local isolates of B. thuringiensis Auky Island Padaido District in Biak Numfor Regency with NCBI gene bank base, the basis of which is to obtain B. thuringiensis isolates from jayapura local isolates that can act as controllers of Anopheles mosquito larvae. Several steps in the research are 16s gene amplification, PCR product purification, cloning using pTA2 vectors and transformation into competent E. coli Zymo 5α cells, confirmation with PCR colonies, recombinant plasmid isolation, sequencing analysis and phylogenetic tree construction. The isolates of ABNP8, ABNP9, ABNP11, ABNP12 and ABNP18 have been detected as local isolates from in Auky Island Padaido District in Biak Numfor Papua Regency that have great potential as bioinsecticides, and capable of controlling and killing Anopheles mosquito larvae. Of the five isolates, ABNP8 isolates had unique diversity and characteristics and were different from the four other isolates. Based on the similarity analysis in the MEGA7 program, the similarity rate reached 84%. Its diversity can be seen from the uniqueness of the sequence and its position in different branching dendrograms.

scholarly journals Characterization of the Thermal Stress Response ofCampylobacter jejuni

1998 ◽  
Vol 66 (8) ◽  
pp. 3666-3672 ◽  
Author(s):  
Michael E. Konkel ◽  
Bong J. Kim ◽  
John D. Klena ◽  
Colin R. Young ◽  
Richard Ziprin
Keyword(s):  
Heat Shock ◽  
Heat Shock Proteins ◽  
Recombinant Plasmid ◽  
Gastrointestinal Disease ◽  
Open Reading Frame ◽  
Calculated Molecular Mass ◽  
Reading Frame ◽  
E Coli ◽  
Shock Proteins

ABSTRACT Campylobacter jejuni, a microaerophilic, gram-negative bacterium, is a common cause of gastrointestinal disease in humans. Heat shock proteins are a group of highly conserved, coregulated proteins that play important roles in enabling organisms to cope with physiological stresses. The primary aim of this study was to characterize the heat shock response of C. jejuni. Twenty-four proteins were preferentially synthesized by C. jejuni immediately following heat shock. Upon immunoscreening ofEscherichia coli transformants harboring aCampylobacter genomic DNA library, one recombinant plasmid that encoded a heat shock protein was isolated. The recombinant plasmid, designated pMEK20, contained an open reading frame of 1,119 bp that was capable of encoding a protein of 372 amino acids with a calculated molecular mass of 41,436 Da. The deduced amino acid sequence of the open reading frame shared similarity with that of DnaJ, which belongs to the Hsp-40 family of molecular chaperones, from a number of bacteria. An E. coli dnaJ mutant was successfully complemented with the pMEK20 recombinant plasmid, as judged by the ability of bacteriophage λ to form plaques, indicating that theC. jejuni gene encoding the 41-kDa protein is a functional homolog of the dnaJ gene from E. coli. The ability of each of two C. jejuni dnaJ mutants to form colonies at 46°C was severely retarded, indicating that DnaJ plays an important role in C. jejuni thermotolerance. Experiments revealed that a C. jejuni DnaJ mutant was unable to colonize newly hatched Leghorn chickens, suggesting that heat shock proteins play a role in vivo.

scholarly journals Effects of Sr2 + on the preparation of Escherchia coli DH5α competent cells and plasmid transformation

PeerJ ◽  
2020 ◽  
Vol 8 ◽  
pp. e9480
Author(s):  
Yonggang Wang ◽  
Xinjian Wang ◽  
Linmiao Yu ◽  
Yuan Tian ◽  
Shaowei Li ◽  
...  
Keyword(s):  
Heat Shock ◽  
Regression Model ◽  
Energy Dispersive Spectrometer ◽  
Regulation Mechanism ◽  
Transformation Mechanism ◽  
Bacterial Gene ◽  
E Coli ◽  
Plasmid Transformation ◽  
Competent Cells ◽  
Cell Surface Structure

Bacterial gene transformation used with Escherichia coli as a desired microorganism is one of the important techniques in genetic engineering. In this study, the preparation of E. coli DH5α competent cells treated with SrCl2 and transformation by heat-shock with pUC19 plasmid was optimized by Response Surface Methodology (RSM). Other five E. coli strains including BL21 (DE3), HB-101, JM109, TOP10 and TG1, three different sizes plasmids (pUC19, pET32a, pPIC9k) were used to verify the protocol, respectively. The transformation mechanism was explored by scanning electron microscope combined with energy dispersive spectrometer (SEM-EDS), atomic absorption spectroscopy (AAS) and Fourier-transform infrared spectroscopy (FT-IR). An equation of regression model was obtained, and the ideal parameters were Sr2 + ions of 90 mM, heat-shock time of 90 s and 9 ng of plasmid. Under this conditions, the transformation efficiency could almost reach to 106 CFU/µg DNA. A small change of the cell surface structure has been observed between E. coli DH5α strain and competent cells by abovementioned spectrum technologies, which implied that a strict regulation mechanism involved in the formation of competent cells and transformation of plasmids. An equation of regression model for the competent cells preparation and plasmid transformation could be applied in gene cloning technology

Evolutionary tuning of heat shock resistance in E. Coli

2020 ◽  
Author(s):  
Joost Schymkowitz ◽  
Frederic Rousseau ◽  
Abram Aertsen ◽  
Bert Houben ◽  
Sebastien Carpentier ◽  
...  
Keyword(s):  
Heat Shock ◽  
E Coli ◽  
Shock Resistance ◽  
Evolutionary Tuning

In Vitro Holdase Activity of E. coli Small Heat-Shock Proteins IbpA, IbpB and IbpAB: A Biophysical Study with Some Unconventional Techniques

2014 ◽  
Vol 21 (6) ◽  
pp. 564-571 ◽  
Author(s):  
Sourav Roy ◽  
Monobesh Patra ◽  
Suman Nandy ◽  
Milon Banik ◽  
Rakhi Dasgupta ◽  
...  
Keyword(s):  
Heat Shock ◽  
Heat Shock Proteins ◽  
Small Heat Shock Proteins ◽  
E Coli ◽  
Biophysical Study ◽  
Shock Proteins

The Small Heat-shock Proteins IbpA and IbpB in E. coli: The Small Ones With a Big Role

2016 ◽  
Vol 9 (2) ◽  
pp. 84-96
Author(s):  
Sanchari Bhattacharjee ◽  
Rakhi Dasgupta ◽  
Angshuman Bagchi
Keyword(s):  
Heat Shock ◽  
Heat Shock Proteins ◽  
Small Heat Shock Proteins ◽  
E Coli ◽  
Shock Proteins

scholarly journals Development and comparison of cell-free protein synthesis systems derived from typical bacterial chassis

2021 ◽  
Vol 8 (1) ◽  
Author(s):  
Liyuan Zhang ◽  
Xiaomei Lin ◽  
Ting Wang ◽  
Wei Guo ◽  
Yuan Lu
Keyword(s):  
Protein Synthesis ◽  
Protein Production ◽  
Influential Factors ◽  
Competent Cell ◽  
Free Protein ◽  
Application Range ◽  
E Coli ◽  
Short Time ◽  
Cell Free Protein Synthesis

AbstractCell-free protein synthesis (CFPS) systems have become an ideal choice for pathway prototyping, protein production, and biosensing, due to their high controllability, tolerance, stability, and ability to produce proteins in a short time. At present, the widely used CFPS systems are mainly based on Escherichia coli strain. Bacillus subtilis, Corynebacterium glutamate, and Vibrio natriegens are potential chassis cells for many biotechnological applications with their respective characteristics. Therefore, to expand the platform of the CFPS systems and options for protein production, four prokaryotes, E. coli, B. subtilis, C. glutamate, and V. natriegens were selected as host organisms to construct the CFPS systems and be compared. Moreover, the process parameters of the CFPS system were optimized, including the codon usage, plasmid synthesis competent cell selection, plasmid concentration, ribosomal binding site (RBS), and CFPS system reagent components. By optimizing and comparing the main influencing factors of different CFPS systems, the systems can be optimized directly for the most influential factors to further improve the protein yield of the systems. In addition, to demonstrate the applicability of the CFPS systems, it was proved that the four CFPS systems all had the potential to produce therapeutic proteins, and they could produce the receptor-binding domain (RBD) protein of SARS-CoV-2 with functional activity. They not only could expand the potential options for in vitro protein production, but also could increase the application range of the system by expanding the cell-free protein synthesis platform.

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