Many consider the twentieth century the "century of the atom" due to the many innovations made possible by new understandings of the elements' physical and chemical properties. These included, but are not limited to, the development of atomic energy (and weapons), diagnostics for medicine, computers, and the microchip, among others.
These breakthroughs shaped the potential and future of science and technology and profoundly impacted our daily lives. But we're living through what may be dubbed the "century of DNA." DNA is like the soundtrack to life's complex symphony; it serves as a template that dictates how cells work. Recent decades have seen tremendous progress in reading and synthesizing DNA sequences, greatly enhancing our capacity to understand and manipulate biological systems.
In today's post, we will consider two major branches of biological engineering: gene synthesis and traditional cloning. This blog will review the main benefits and drawbacks of each method to assist you in choosing the appropriate approach for your project.
Let's begin with a closer look at the two approaches.
What is Gene Synthesis?
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Gene synthesis is the in vitro method of producing a gene without requiring a preliminary template.
Alternatively, if you want to have a more in-depth explanation, synthetic gene creation is a technique that involves a series of steps for producing a gene and other gene derivatives without the need for a DNA template. As a result, it streamlines the process of making various genes, notably custom genes with unique base pairs and sequences.
Har Gobind Khorana and colleagues performed the first effective gene synthesis using a yeast tRNA in 1972. Since 1972, advances in gene assembly and molecular biology techniques have allowed the successful construction of genetic sequences unique to a broad range of scientific applications.
Steps Involved in Gene Synthesis
A general overview of the steps toward a successful gene synthesis includes the following:
- Oligonucleotide synthesis
- Oligonucleotide annealing
- Cloning of gene sequences
- Screening for clones
- Analyzing sequences and correcting mistakes
A couple of decades ago, the idea of using gene synthesis for targeted gene therapy or other advanced applications would have seemed entirely out of reach. Today, gene synthesis opens up a world of possibilities that go far beyond what was previously imaginable, with applications ranging from gene regulation to understanding evolution and antibiotic resistance.
Thanks to advances in genetic engineering, researchers can now construct custom antibodies, variant libraries, custom peptides, operons, and proteins with improved or novel functions.
Now, let's shift our focus to the more traditional method of doing things.
What is Traditional Cloning?
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Traditional cloning is a method of DNA manipulation that typically involves using restriction endonucleases to generate DNA fragments with specific complementary ends. These fragments are then joined together using a DNA ligase and introduced into a host organism through transformation.
To do this, you usually need to cut the DNA sequence in two places so that there are two restriction sites on either side of the DNA sequence. These sites also correspond to the best place for the vector for sequence insertion, sometimes called the multiple cloning site (MCS).
By applying two separate restriction enzymes (REs), incompatible ends are produced, which allows for directional cloning of the insert and helps reduce the transformation background caused by the re-ligated vector.
Pros and Cons of Traditional Cloning
Pros
- Low costs
- Flexible
- A wide variety of vector options
- Performing directional replication is easy and stress-free
Cons
- A restriction site's existence and its effect on translation may impose sequence limitations.
The first step in conventional cloning using recombinant DNA technology is to use restriction enzymes to prepare a vector to accept an insert DNA. Following digestion, the fragments are joined by the enzyme DNA ligase, a procedure known as ligation, which creates a new vector that can express a desired gene.
This is the most basic and traditional method of genetic material production, and it served as the basis for the development of novel cloning techniques that capitalize on the unique properties of other modifying enzymes, including:
- TA cloning,
- TOPO cloning,
- PCR cloning,
- Ligation-independent cloning, and
- gene assembly.
You can summarize the steps involved in typical traditional cloning as:
- Vector preparation
- Insert preparation
- Ligation
- Transformation
- Colony screening
Given this information, let's compare the two (gene synthesis vs traditional cloning) to better understand their differences.
Traditional Cloning vs. Gene Synthesis: Major Differences
When compared to molecular cloning and polymerase chain reaction (PCR), two of the more conventional approaches to gene cloning, gene synthesis has numerous significant benefits. There are two main limitations to these traditional approaches. First, they can only amplify and clone genes; they cannot synthesize them.
Another limitation of restriction-based molecular cloning and PCR is that the genetic sequence of interest must first be acquired and then amplified. This is a prerequisite for both techniques. With gene synthesis technology, the desired genetic sequence can be created and amplified without physically obtaining it, which is a significant upside.
Current gene expression technologies enable the synthesis of de novo sequences. This allows for the generation of sequences of interest even without physical copies of genes, such as viral DNA or cancer cell mutations in particular genes. The lack of a physical sample of the target gene considerably reduces the financial and time investments required when compared to conventional approaches such as cloning, amplification, gene knockouts, and targeted mutagenesis.
Owing to these benefits, gene synthesis has grown in popularity across many scientific domains because of its user-friendliness and cost-effective workflow.
Suffice it to say that both techniques have unique applications based on the requirements of a given project.
Continue reading to learn some essential applications of these two approaches.
Applications of Gene Synthesis and Traditional Cloning
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The Uses of Gene Synthesis in Real Life
1. Vaccine Research And Development
In vaccine research, gene synthesis is widely employed in everything from fundamental immunology and microbiology studies that are essential for rational vaccine design to state-of-the-art DNA synthesis and vaccine development initiatives. The development of new vaccines is necessary to combat both newly emergent threats, such as the Ebola virus, and persistent infectious viruses, such as the seasonal influenza A virus.
Meanwhile, cancer vaccines have twofold purposes: first, as a preventative measure against oncogenic virus infection; and second, as a therapeutic tool to mount an immune response against malignancies.
Building codon-optimized custom genes encoding adjuvants, customized vectors tailored for safety, and gene cassettes that control the expression of specific antigenic proteins are all possible using gene synthesis.
2. Plant Biology Research
Gene synthesis can be used in plant biology research to create DNA vaccines by designing gene cassettes that encode specific antigenic proteins, constructing codon-optimized synthetic genes that produce adjuvants, and developing safety-optimized vectors.
3. Therapeutic Antibodies
It is possible to activate, suppress, or modify endogenous immune responses to particular cells or molecules with therapeutic antibodies, particularly monoclonal antibodies. The treatment of numerous diseases, including cancer, inflammatory and autoimmune disorders, and many others, is being transformed by antibody-based medications. Some examples include:
- Therapeutic Inhibition of Inflammation in Pigs and Humans by Chimeric Anti-CD14 IGG2/4 Hybrid Antibodies.
- In experimental autoimmune encephalomyelitis in mice, removing autoantibodies improves the symptoms.
- One Human Monoclonal Antibody Cross-Neutralizes Four Paramyxoviruses.
4. Brain research
A lot of neuroscience research uses gene synthesis. For example, codon-optimized channelrhodopsins make it possible to use optogenetic methods to see how neurons are connected; customized ORF clones and RNAi constructs help to understand how glioblastoma cells communicate; and gene variants are used to look into how neurons communicate and change how they connect and learn, to name a few examples.
The Uses of Traditional Cloning in Real Life
- The traditional cloning of genes has empowered the development of proteins, medicines, and other products. These products can take many forms, including cloned DNA, cells, tissues, or animals.
- When it comes to agriculture, cloning is utilized to develop pest-resistant crops.
- In addition, cloning is used for gene therapy and the creation of transgenic animals.
- The process of cloning cattle allows for the duplication of desirable traits, such as fast growth and high milk output, without the genetic "lottery" and endless blending during sexual reproduction.
- It allows for the faster breeding of an animal with a specific genetic modification compared to regular mating, for example, the ability to produce medication in its milk.
- Furthermore, modifying a gene in cultured cells is much simpler than doing so in an entire animal. Moreover, combining the altered cell nucleus with an enucleated egg makes a clone of the desired genotype possible.
- Animals used in experiments, like mice, are cloned so researchers may learn more about their fundamental biological processes.
Choosing Between Gene Synthesis Services And Traditional Cloning Services
Although numerous custom gene production services exist in the market, the right one usually offers supplementary benefits that extend beyond the DNA itself.
Here are some things you should look for when choosing gene synthesis services.
Dependability: Over 21 years of experience in the synthesis of synthetic DNA and a 5-star Google review, dependable 100% sequence-verified genes assured by ISO-accredited quality systems, and environmentally responsible production practices.
Enhanced efficiency and economy of scale: With the right exclusive program, you can improve your sequence for dependable and improved expression in your host system while reducing the cost per milligram of protein.
Stellar customer experience: Placing an order for synthetic biology products has never been easier. It's now possible to request a quote at any time and from any location at the click of a button. You won't have to wait days for an estimate or optimization results. The only thing you need to do is submit your sequence, choose your criteria, and then place your order. Any questions you may have about gene synthesis can be quickly addressed by contacting our knowledgeable customer service team.
What you require as a product: Depending on your requirements and budget, you may choose between a variety of custom genes, antibodies, or custom peptides from our product lineup, or you can choose to create your project from the ground up. We offer a range of products, from DNA fragments to expression-ready designs, DNA libraries, and protein expression and purification services. Everything is handled internally by our experts.
High Success Rate: Our proficiency in custom gene synthesis is substantiated by the fact that we excel in producing genes with multiple complexities while accomplishing exceedingly challenging sequences with a success rate of 99%.
Variety of Platforms: Companies that exclusively depend on silicon-based platforms are limited to developing simple and short genes, which can hinder their capacity to work on a significant portion of gene projects up to 50%.
Our accomplishments may be attributed to more than two decades of experience, during which we have created and employed special reagents and enzymes in our manufacturing procedures, all while following the most reliable synthesis techniques.
Summary
Commercial gene synthesis services are rapidly becoming a viable, less expensive substitute for conventional molecular biology methods like cloning, which goes against the grain of what many researchers previously believed. The primary reasons are as follows:
- Reduced workload - The conventional approach to cloning is a multi-step procedure that includes the design of a cloning strategy, primer production, polymerase chain reaction (PCR), gel extraction, and bacterial transformation, among other sophisticated steps. Unlike gene synthesis, these procedures demand a substantial investment of time and human resources.
- Minimizes expenses - Most of the time, ordering a manufactured gene is cheaper than ordering cloning kits, oligos, or DNA/gene sequencing services.
- Improved DNA performance - Codon optimization is possible during gene synthesis and has been shown to make protein translation more efficient.
- Practicality - Researchers can get any gene they want by simply providing the nucleotide or amino acid sequence. This is because they don't need a physical template and aren't limited by design constraints that come with traditional cloning.
Irrespective of which camp (gene synthesis or traditional cloning) you decide to join, you'll need reliable, high-quality supplies like ELISA kits, bioreagents, and peptides. Luckily, Biomatik is a one-stop shop for these biochemical products, where you can find services like custom gene synthesis, recombinant protein production, and more. Reach out today!