Biogenius: Gene Cloning and Expression [Theory & Practice]

Name: Biogenius: Gene Cloning and Expression [Theory & Practice]
Rating: 4.7 (19 reviews)

"Animated Theory, Simulated Practice, Guaranteed Results: 30-day Money Back!"

Created byOmar Taha

Last updated 2/2024

English

What you'll learn

Understand the foundational principles of recombinant DNA technology and its historical significance.
Master the ligation process essential for joining DNA fragments in gene cloning.
Acquire proficiency in performing restriction digestion experiments and manipulating DNA fragments.
Grasp the transition of plasmids into cloning vectors and the importance of vector selection.
Examine the pioneering work of Cohen and Boyer in creating the first plasmid vector and how these vectors work.
Analyze the structure and significance of the pBR322 plasmid in genetic engineering
Develop effective cloning strategies and troubleshoot common challenges in gene cloning
Explore the characteristics and applications of PUC series plasmids
Master the construction and utilization of DNA libraries in lambda phage for gene identification.
Understand the role and applications of M13 phage vectors in DNA sequencing.
Explore the features and applications of DNA libraries in COSMIDs for improved gene cloning.
Learn the principles behind constructing cDNA libraries and their applications in gene expression.
Master the PCR technique for efficient DNA amplification.
Understand the principles and applications of TA cloning for simplified gene insertion.
Gain a comprehensive understanding of the lac operon and its role in gene regulation.
Explore strategies for modifying the lac operon to suit specific research objectives.
Identify and comprehend key genetic elements crucial for gene expression.
Understand the significance of RBS, start and stop codons in translation and protein synthesis.
Explore different promoters, including P lac and P Tac, and their applications in gene expression.
Investigate host-specific promoters such as Pbad and Rha Pbad in gene regulation.
Explore the use of phage promoters in gene expression and their advantages and challenges.
Understand the two-step system for controlled gene expression and promoter combinations.
Grasp the role of replication origin in plasmid replication.
Master the technique of co-expressing genes from two plasmids.
Differentiate between various types of expression vectors and understand gene insertion principles.
Evaluate the advantages and limitations of bacterial expression systems.
Define the objectives and diverse applications of gene expression studies
Understand the concept of expressing fusion proteins and their applications.
Explore pull-down assays for studying protein interactions and understand signal peptides' role.
Master the principles and applications of Topo cloning for simplified gene cloning.
Identify and characterize expression strains and understand their contribution to efficient gene expression.

Course content

3 sections • 39 lectures • 6h 27m total length

Introduction to recombinant technology5:03
Ligation reaction8:29
Discover how the ligase enzyme joins DNA fragments at the five-prime and three-prime ends through dehydration synthesis, driven by ATP converting to AMP and enabling nucleophilic attack.
Restriction digestion14:31
From plasmid to cloning vector10:54
Cohen and Boyer: the first plasmid vector1:39
pBR322 plasmid7:49
Cloning strategy2:57
PUC series plasmids8:34
DNA libraries in lambda phage26:37
M13 phage6:07
Libraries in COSMIDs5:17
Explore DNA libraries and cosmids, using cos sites and cosmid vectors to assemble genomic fragments into concatemers, package into bacteriophages, and select ampicillin-resistant colonies in E. coli.
cDNA library16:15
PCR13:42
TA cloning4:24

Understanding Lac operon11:43
Modifying Lac operon5:24
Genetic Elements Essential for Expression6:30
Explore expression plasmids by configuring a promoter, operator, ribosome binding site, start and stop codons, terminator, a multiple cloning site, and an origin of replication with a selectable marker.
RBS START and STOP codons2:20
Promoters From P lac to P Tac11:09
Host promoters: Pbad and Rha Pbad12:54
Phage Promoters4:35
Two-step system and promoter combinations3:26
Demonstrate a two-step system by combining T7 promoter with lac or pbad promoter on a plasmid, inducing T7 polymerase via IPTG or arabinose to drive high-level expression.
Replication Origin1:52
Learn how replication origin in plasmids determines copy number, with pUC from the University of California yielding the highest copies, about 500–700 per bacterial cell, unlike pac1 plasmids.
Co-expression from two plasmids1:14
Explore co-expression from two plasmids in the same bacterial cell, producing two different proteins, and its use in yeast two-hybrid systems to study protein-protein interactions with bait and prey.
Types of expression vectors: Insertion into vectors12:42
Pros and cons of Bacterial expression systems13:54
Explore the pros of bacterial expression systems—fast growth, cheap media, and simple host biology. Understand cons like codon bias, limited post-translational modifications, and endotoxin risks requiring careful purification.
Goals of expression6:44
Explore the goals of expression: maximize soluble, properly folded protein while preserving cell viability. Adjust inducer level, timing, and temperature to reduce aggregation and aid purification.
Expression of Fusion proteins11:55
Explore expression and purification of fusion proteins using his-tag and GST-tag strategies, including affinity chromatography with Ni-NTA, protease cleavage, IPTG induction, and MCS placement.
Pull-down assays and signal peptides7:38
Topo Cloning9:31
Expression strains10:49

Study objective, GFP uses, example of Biosensor6:38
Scheme of work4:29
Total RNA extraction19:41
Extract total RNA from jellyfish with Trizol, containing guanidinium thiocyanate and phenol, perform di phasic separation with chloroform, precipitate with isopropanol, wash with ethanol, and elute in a T buffer.
Quantify RNA (using Nanodrop)8:08
cDNA synthesis25:32
Get the sequence of gene insert11:05
Primer design17:00
Primer design: Which REs should we use?27:53

Requirements

No prerequisites needed

Description

This cutting-edge course is meticulously crafted to empower aspiring molecular biologists with the essential knowledge underpinning recombinant DNA technology and genetic manipulation. As a participant, you will embark on a journey that transcends theoretical complexities, providing you with a robust foundation for engaging in practical, real-world applications.

The curriculum spans a wide spectrum of topics, ranging from the fundamental principles of molecular biology to the intricacies of gene cloning techniques. Through a blend of in-depth lectures, interactive discussions, and case studies, you will not only grasp the theoretical intricacies but also cultivate the analytical skills necessary to tackle multifaceted challenges.

Furthermore, this course places a strong emphasis on hands-on experiences, ensuring that you are well-prepared for the dynamic landscape of molecular biology. Laboratories and workshops will offer you the opportunity to apply theoretical knowledge in a practical setting, fostering a seamless transition from theory to application.

As you progress, you will explore advanced topics such as expression systems, recombinant DNA technology, and ethical considerations in genetic engineering. The goal is not only to deepen your understanding but also to equip you with the tools and insights needed to contribute meaningfully to the rapidly evolving field of biotechnology.

By the end of this transformative program, you will emerge not only with a profound theoretical grasp of molecular biology but also with practical expertise, ready to shape the future of biotechnology and molecular research. Join us on this educational odyssey, and become a proficient molecular biologist prepared to make a significant impact in the ever-evolving world of genetic science.

Who this course is for:

Entrepreneurs in Biotech Startups
Biochemistry Graduates
Undergraduate Biology Students with Research Aspirations
Healthcare Professionals
Veterinary Scientists
Pharmacologists
Medical Students
Microbiologists
Medical Laboratory Technologists
Neuroscientists
Botanists
Bioinformatics Specialists
Aquatic Biologists
Food Scientists and Technologists
Immunologists
Clinical Researchers
Biological Anthropologists

Biogenius: Gene Cloning and Expression [Theory & Practice]

What you'll learn

Explore related topics

Course content

Theoretical: Cloning14 lectures • 2hr 12min

Theoretical: Expression17 lectures • 2hr 14min

Practical8 lectures • 2hr

Requirements

Description

Who this course is for: