Molecular Biology - Synbio Technologies

Tag: Molecular Biology

Gene Synthesis Plasmid Preparation

At Synbio Technologies we pride ourselves in being one of the premier companies within not only the gene synthesis industry, but the biotechnology industry as a whole. This confidence is relying upon our tested and proven methods of gene synthesis. Gene synthesis can be defined as the method in synthetic biology used to engineer an artificial gene of interest in a laboratory setting. Ever since gene synthesis was first successfully conducted in the early 1970s it has become a highly sought mechanism in various fields of genetics research. For this reason many companies, like Synbio Technologies, have been attempting to optimize their gene synthesis products; but none of them have done this with quite success like Synbio Technologies. Our Syno Platform allows us to generate any sequence of interest up to and including 200kb in length. Another aspect of gene synthesis that Synbio Technologies has to offer is our gene synthesis plasmid preparation. Using this technology we are capable of generating any sequence and inserting it into any plasmid of interest. For gene synthesis plasmid preparation there are typically two forms of final product: transfection grade and research grade. Both differ slightly in their mechanism of being generated, as well as research application. Through use of Synbio Technology’s Syno^®Platform, as well as our gene synthesis plasmid preparation we are ready to supply our customers with whichever plasmid product is necessary.

The differences between the transfection grade and research grade generated plasmids lies mainly in the applications each particular plasmid is used for. First, the research grade is used more in a laboratory setting to conduct various types of genetics research. These topics include: molecular cloning, mutagenesis, southern blotting, etc. All of these methods are used in order to conduct various types of genetics research with use of gene synthesis. Where gene synthesis comes in is through the insertion of the requested sequence into the plasmid. This insertion, and subsequent amplification, allows our customers to conduct any type of research necessary that requires a research grade plasmid construction. Second, the transfection grade plasmid is used mainly for protein manufacturing, antibody production and other forms of gene therapy. The link between this plasmid and gene synthesis is again the insertion of the sequence of interest into the particular plasmid. This can be extremely useful when conducting different types of gene therapy. The requested sequence can be loaded into the plasmid with the hopes of reversing the endogenous mutated gene of interest. This powerful technology has the ability to further our understanding of genetics and it is all thanks to gene synthesis.

Gene synthesis is something that Synbio Technologies does extremely well, mainly relying upon our Syno^®Gene Synthesis Platform. This platform is used to generate the customer requested sequence with one hundred percent accuracy. The resulting gene synthesis product can then be used in order to prepare a plasmid of interest. This process is normally quite daunting, but Synbio Technologies is more than confident in our ability to generate the requested gene synthesis product and resulting gene synthesis plasmid. This process is all done within a quick turnaround time and competitive prices. For this reason, Synbio Technologies has risen to the top of not only the gene synthesis industry, but the resulting biotechnology industry as well. We offer a one stop shop for our customer’s gene synthesis plasmid preparation, with competitive prices, high quality output, and an efficient turnaround time. With this combination, our customers will be conducting research in no time with confidence in the product that we supply them.

Gene Synthesis Related Services

Synbio Tenchologies can also design sequencing with codon optimization software -NG^TMCodon Optimization Technology at no cost.

Molecular Biology Evolution

Since molecular biology was first established, in the 1930s, the complexity of various biological systems have been explored in order to better understand these systems. As years went by, and our understanding of these systems increased, the information that fell under the, large and ever-growing, umbrella of “molecular biology” became more specific and eventually subdivided by the particular sub-field being studied. Some of these sub-fields include: genetics, proteomics, and cytology. Many of these sub-fields have become extremely popular and very well studied since molecular biology was first established.

Genome Project and Molecular Biology

In the more recent past, there has been an influx of quantitative research within molecular biology. This groundbreaking research has completed the connection between two popular sub-fields of molecular biology: computer science through use of bioinformatics and computational biology. This was most notably seen with the success of the Human Genome Project (HGP). The Human Genome Project allowed for over three billion nucleotide base pairs of euchromatic genome to be sequenced and presented as a reference. This was an important milestone of molecular biology which allowed the human genome to go from the analog world of biology into the digital world. This was a giant step, within the field of molecular biology, which caused a domino effect that resulted in thousands of human genomes to be sequenced with increasingly lower cost.

On 2^rd June 2016, the Genome Project-Write (GP-write) announced a continuation of genomics research through multiple molecular biology tools. These multiple molecular biology tools included gene synthesis and genome editing technologies. These technologies are utilized to synthesize and test large portions of many genomes stemming from microbes, plants and even animals. GP-write’s commitment to furthering our understanding and effectiveness of these technologies has the possibility to improve research and development of the topics of life sciences, new bio-based therapies, and nutrition.

New era of molecular biology

Even as these recent accomplishments in molecular biology unfold, a new problem seems to be stemming from them. The problem that has come about comes in the form of designing and even artificially synthesizing new life.

A recent study has proven the ability to accomplish a complete chromosomal transplantation from one cell to another. After the transplant has been successfully conducted, the chromosome can then be activated to conduct various genetic activity. We can then utilize specific enzymes, digestive proteins and other substances within these cells. This combination will result in the cell’s loss of original features and a totally new species.

Due to the crisis of resource shortage critical to human sustainability partnered with the ever-increasing human population, there is a need for us to seek effective approaches for sustainable living. The furthering of our knowledge on biological systems through these technologies would have many positive effects on successfully creating a sustainable habitat. These positive effects would come as a result of better understanding of the physiology of cells, developing new molecular medicines, as well as generating sustainable energy sources, such as biofuels. All of these will contribute to a more successful living environment through the use of molecular biology.

One molecular biology technology, gene synthesis, has slowly become better understood over the past ten years and has drastically increased our capability of editing and synthesizing genes of interest. Synthesizing DNA artificially is very difficult and increases in difficulty when attempting to synthesize long genetic sequences. This is because the longer the sequence being generated is, the higher the possibility of generating errors. Therefore, a new method is required to successfully conduct gene synthesis and correct for all mistakes generated when the sequence was being synthesized.

As one of the leading companies in the field of synthetic biology, Synbio Technologies has unique proprietary GPS platform on the basis of genotype, phenotype and synotype. We can provide excellent molecular biology services including: plasmid DNA preparation, PCR cloning and subcloning, site-directed mutagenesis and vector construction. We have the ability to generate sequences that are de novo, meaning that the genetic sequence is not preexisting within any organism in biology. We also have the ability to generate sequences up to and including 200kb in length in addition to complex gene products and structures. Synbio Technologies prides itself in our ability to use these molecular biology technologies to better suite our customer’s needs when conducting various types of research.

Molecular Biology Related Services

Molecular Biology Cloning

Molecular biology cloning

Molecular cloning is one method in molecular biology that is commonly used to amplify a genetic sequence of interest. This is accomplished by inserting recombinant DNA into a vector which can then carry DNA fragments in host organisms to be amplified. This process of amplification is based on molecular biology standard, first is to recombine the target gene into the vector DNA molecules in vitro. Then transfer the recombinant DNA to host cells. After transferring, there is a screening of cells which have expressed the recombinant DNA, after purification and amplification.

Molecular Biology Cloning Technology Process:

Isolate the target gene and vector:

1.Direct separation is suitable for the extraction and separation of bacterial chromosomes, plasmids and virus DNA whose genetic background are of interest to be studied.
2.Gene synthesis is used to generate short DNA fragments whose sequence is known clearly.
3.cDNA can be synthesized by reverse transcription from mRNA.
4.Screening the gene of from the genomic library for molecular cloning.

The target gene and vector are cleaved with a restriction enzyme.

This allows the fragments to be more easily connected later.

The target gene and vector are then ligated with DNA ligase.

This seals the connection between target gene and vector.

Transfer the ligated recombinant vector into host cells

Bacteria: E. coli, fungi: Yeast, insect cells or mammalian cells.

Conduct screening at different levels using different methods to test for quality.

For example: vector size, enzyme digestion results, screening markers and so on.

Molecular biology cloning generally uses DNA sequences from two different organisms. First is the species that is the source of the DNA to be cloned. Second is the species that will serve as the living host for replication of the recombinant DNA. Molecular cloning technology is central to many contemporary areas of modern biology and medicine.

Molecular Biology Related Services

Introduction to Molecular Biology

Introduction to molecular biology

Molecular biology is a subfield in biology that studies the various topics and molecular mechanisms within cells that are important for proper cellular function. These topics can include any cellular mechanism ranging from cellular metabolism to utilizing a molecular function in order to successfully accomplish gene synthesis. These topics and mechanisms have the ability to be altered slightly to accomplish a particular application of interest. These mechanisms within molecular biology, have been studied intently and applied to various fields of biological research and development. The applications within molecular biology that this article will introduce are gene expression and protein synthesis. These endogenous mechanisms are vital to the proper cellular function within molecular biology that can be altered to successfully conduct both gene synthesis and protein synthesis.

Molecular biology for gene synthesis

One important aspect of the gene synthesis process is that it is not limited to DNA replication of a preexisting DNA fragment. Gene synthesis is capable of in vitro artificial DNA sequence synthesis, which allows for the synthesis of de novo genetic sequences. Contrary to traditional methods of DNA amplification, gene synthesis does not require that the sequence of interest be preexisting within nature. This is accomplished by using refined chemical methods for gene synthesis without a template DNA chain. The resulting templates are then connected to form a gene fragment, by use of various molecular biology mechanisms. With the development of molecular biology, researchers have the ability to synthesize any gene of interest up to and including 200kb in length. The development of more efficient gene synthesis technologies has also allowed the gene synthesis process to become more time efficient, accurate and cost effective. These contributing factors has led gene synthesis to become an important technique within molecular biology.

Molecular biology for protein express

The aim of gene fragment synthesis is for the resulting synthesized gene to be inserted into plasmid vector. After this is accomplished, the recombinant plasmid transcripts are then inserted into cells, where the recombinant plasmid will be expressed. This allows for the expression of the newly synthesized gene of interest and its resulting protein. In order to obtain the highest quality product and strongest protein expression the recombinant protein must first be purified. The resulting purified protein can then be used in subsequent experiments, specific to our customer’s interests. These experiments may include research contributing to the better understanding of protein structure, function and activity of cells, as well as the diagnosis of the certain diseases. All of applications can provide scientific basis for treatment and drug development. For these reasons, optimizing the gene synthesis expression at the protein level is of upmost important to us, at Synbio Technologies.

Synbio Technologies offers comprehensive molecular biology services, designed to fit our customer’s specific needs. These molecular biology services include gene synthesis, PCR cloning, subcloning, site-directed mutagenesis, vector construction and other related projects. The services Synbio Technologies offers rely upon our Syno® 1.0 and Syno® 2.0 platforms. These platforms allow us to generate the highest quality gene synthesis product for a cost effective price. In addition to this, Synbio Technologies also offers the bacterial and yeast expression systems. These systems help optimize the resulting protein product from our synthesized sequences. The whole process is done with such ease and efficiency, that we are confident our high quality purified recombinant protein will arrive to your bench in as soon as 6 weeks.

Molecular Biology Related Services

Applications of Molecular Biology Techniques in Environmental Microbiology Research

Advances in molecular biology and genetic engineering technology, microbial genetic manipulations have promoted the application of microorganisms in in ecological and environmental research. Genetically engineered microorganisms are being developed and assessed for their beneficial uses in environmental monitoring, toxic chemicals pollution control and genetically engineered microorganisms.

Hybridization probe

The oligonucleotide probes can hybridize to DNA or RNA whose base sequence allows probe–target base pairing due to complementarity between the probe and target to analyze the presence of nucleotide sequences (the DNA target) that are complementary to the sequence in the probe.

Due to the stringent specificity and high sensitivity of nucleic acid hybridization, hybridization probes are used on a broad level in microbial ecology, such as microbial detection, qualitative and quantitative analysis of microbial, distribution, abundance and adaptability of microbial.

PCR based technologies

The polymerase chain reaction (PCR) is a technique used in molecular biology to amplify DNA template, generating thousands to millions of copies of a particular DNA sequence in vitro. This technique can be used to analyze mRNA expression profile among different growth stages.

Electrophoresis

The interaction between DNA double helix are disrupted during denaturing gradient gel electrophoresis(DGGE)， temperature gradient gel electrophoresis(TGGE) and other special forms of electrophoresis, thus the DNA fragments consist of different sequences can be separated on acrylamide gel with superior resolution.

Genetic engineering

New recombinant DNA may be generated by first isolation and amplification the genetic material of interest using molecular cloning methods, then the chimera DNA sequence or artificially synthesized DNA maybe inserted into the host organism. This technique is essential for the construction of microbes with enhanced biodegradability that may be used in controlling remediation of contaminated environment or fermentation of waste to produce natural gas.

Application of molecular biology technology not only expanded the horizon but also increased the depth of microbial ecology research. The increasing amount of microbe’s genomic data provides new opportunities for understanding the genetic and molecular bases of the degradation genes in various bacteria. The in-depth knowledge of microbes’ genome will make the research more objective and more controllable.

Applications of PCR Cloning Technology

PCR cloning technology can amplify trace amounts of DNA, generating millions of copies of a specific DNA sequence. PCR is highly sensitive, extremely specific, and high-yield, while also being easily reproducible, fast, and convenient, making it an amazingly powerful tool in molecular biology. With the rapid development of modern life science, PCR cloning technology has been more and more widely applied to various fields in biological research, medical research, virus detection, and the food industry.

PCR cloning Technology Applied to Gene Cloning

Gene cloning and Subcloning via PCR technology plays a significant role in cell biology research. PCR technology can generate millions of copies of a single-copy gene, amplifying a specific DNA fragment that might only be a few picograms. Compared to other gene cloning techniques, PCR omits several tedious processes for cloning of a particular gene fragment from genome DNA, such as enzyme digestion, connection, transformation, DNA library construction, gene screening, gene identification, and Subcloning.

PCR Cloning Technology Applied to DNA Recombination

In molecular biology, PCR cloning technology can be used to construct recombinant DNA molecules by inserting different sources of specific genes or DNA fragments into viruses, plasmids or other vectors in vitro. Recombinant DNA molecules are then imported into reporter cells to amplify and reproduce. After screening, the daughter cells that contain the target gene are further multiply to extract a large amount of DNA. Recombinant DNA technology can be applied to the Human Gene Project, valuable protein expression, gene diagnosis and therapy, genetic modification of animals and plants, and other research fields.

PCR Cloning Technology for Gene Quantification

PCR cloning technology can be applied to quantitatively determine the copy number of a target gene in a sample. The target gene and a single copy reference gene are placed in a tube for PCR cloning. The PCR product is then separated by electrophoretic separation technology and band intensity is observed. Alternatively, the 5’ end of the primer can be marked by a radionucleotide, after which the gene copy number can be determined by radioactivity quantification. PCR cloning technology can also be applied to quantitatively analyze mRNA and tRNA. It can even detect 1TIRNA, which is hard to detect even by Northern blot.

Alteration of endogenous genes and invasion of foreign genes can be threatening to human health. Regardless of whether or not pathogenesis is caused by genetic changes, as long as there is a pathogen, its corresponding existing nucleic acid can be found. With the development of PCR cloning technology and related technologies, PCR can be applied to infectious disease pathogenesis detection and diagnosis, tumor related gene detection, hereditary disease early diagnosis, bone marrow transplant HLA – D locus matches, and evolutionary theory analysis.

Synbio Technologies’s Syno^®2.0 platform can clone a target gene to any specific point on provided vectors without depending on restriction enzyme sites, and can quickly and cost-effectively fulfill a wide variety of client requests for myriad applications in synthetic biology.

The Methods of PCR Cloning

PCR Cloning is a technique used to amplify a specific region of DNA strand, and is used in almost every molecular biology lab in the world. PCR can be used on almost any DNA region, provided that suitable primers can be made. Synbio Tech provides one-stop PCR cloning services, including primer design.

PCR Primer Design Procedure

Acquisition of DNA sequence: For amplification of known DNA sequences, tried-and-tested primers can be found on the NCBI website. For unknown DNA sequences containing conserved sequence(s) from related species, primers should be designed according to the DNA or RNA of the conserved sequence(s).
PCR primer design: Many commercial software products and online tools are used to design primers. Primer Premier 5.0, the most popular primer design software, is both powerful and convenient to use. It can also contrast and comprehensively assess the best choice of primer according to their specificity.
Validation of PCR amplification: Once primer synthesis finishes, the accuracy of the primer can be predicted after gel electrophoresis of PCR product.

Notes About PCR Primer Design

The length of primer should be around 18-24bp. If the primer is too short, the primer specificity will be too low; if the primer is too long, it may cause base pair mismatches and may reduce the PCR amplification efficiency.
The GC% of the primer will affect the denaturation temperature (Tm). Tm should be around 55-80℃ and the annealing temperature difference between the upstream and downstream primers should be within 10℃. Usually, the GC% of primer should be between 40%-60% ,and the GC% difference between the upstream and downstream primers should be within 20% of each other in order to enhance primer specificity.
Repetitive structures or high similarity with the template sequence should be avoided as both may lead to possible base pair mismatches.
Secondary structure of primers may inhibit the PCR reaction, and should be avoided.

Synbio Technologies provide one-stop PCR cloning services, including primer design. Our Syno^®2.0 platform can clone the target gene to any specific point on a provided vector without relying on restriction enzyme sites, to best satisfy the cloning requirements of each client.

PCR Techniques in Biology

Polymerase chain reaction, generally known as PCR and also referred to as in vitro DNA amplification, is one of the most widely used techniques in biology labs across the world.

Traditional DNA amplification method

Traditional DNA amplification involves the construction of a vector containing the target gene, followed by the transfer of the vectors into cells to amplify the target gene and then screening by probe. The process includes enzyme digestion, connection, transformation, culture, and probe hybridization. Although there are no technical difficulties for the traditional method, the complicated operation and long cycle length make it suboptimal as a way to amplify DNA. PCR is faster, more efficient, and cheaper, and has many advantages over traditional DNA amplification.

Advantage of PCR cloning

PCR was first theorized by Kary Mullis in 1983 and was later invented in 1985. PCR can amplify traces of DNA and generate millions of copies of a particular DNA sequence. Because of its high sensitivity, specificity, and yield, along with its reproducibility, speed, and convenience, PCR is now widely used in microbiology, medical science, agriculture, and many other fields. PCR has greatly simplified the process of molecular cloning, enabling researchers to much more easily analyze and identify genes of interest.

Basic principles of PCR cloning

At a temperature of 95℃, DNA denatures and yields single-stranded DNA molecules. The primer then binds to a complementary part of the DNA template. The temperature is then lowered to the optimum activity temperature, which is usually around 72℃. This activates DNA polymerase, which synthesizes a new DNA strand complementary to the DNA template in the 5’ to 3’ direction. The DNA template, primers, and polymerase are then thermocycled through denaturation, annealing, and extension steps, allowing DNA polymerase to replicate a target region of DNA by millions of times or more.

PCR has become an essential part of biology labs around the world, and is widely applied to gene cloning, genetic recombination, DNA sequence analysis, and gene quantification. PCR Cloning is also used in cancer gene detection and early diagnosis of hereditary diseases.

Our Syno^®2.0 platform can clone any target gene to any specific point on required vectors
without depending on restriction enzyme sites. We promise both accurate and speedy delivery of any client’s cloning requests.

Codon Usage Bias

The genetic information in DNA is transcribed into mRNA then translated into protein, in which codons played important roles during the process. Total 64 combinations of nucleotide triplet (codon) encode all 22 amino acids. Each amino acid corresponds to at least one codon, e.g. methionine and tryptophan. In other cases, amino acids are encoded by 2 to 6 different codons. The codons encode the same amino acid are referred as synonymous codons. The frequency of synonymous codon usage varies widely among different organisms, and these differences have important implications for the regulation of protein expression. In the process of protein synthesis, a particular species tends to use a set of specific codons, which are called optimal codons. This phenomenon is also known as codon usage bias. So the use of different species in the codon usage bias, optimizing the use of codons can increase the protein expression.

Codon usage bias and protein

At the same time, accurate polypeptide elongation could minimize the energy waste caused by translation errors. Therefore, a codon with optimal translation speed and high fidelity will result high translation efficiency and an increased protein expression. The combination of codons and translation efficiency are not only contributing to the codon usage bias of the whole genome, but also the distribution of codons with different translation efficiency in different regions. The codon usage bias in different regions can regulate genes expression at different stages, such as affecting the mRNA expression during transcription, the speed and accuracy of translation, and the folding of polypeptide.

In the heterologous protein expression system, selecting the codon combinations that control the speed of translation is essential to avoid formation of inclusion bodies due to high expression or not obtaining any protein at all. Thus a well-tuned system for controlled gene expression is the key component for protein production in industrial and scientific research.

Synbio Tech proprietary NG^TMCodon Optimization software can intelligently optimize codons based on different expression systems and effectively improve the protein expression in order to meet the needs of scientific research and industrial production.

Synbio Technologies’ Codon Optimization Strategy

Translation is the last step in protein synthesis that results in stable proteins from DNA replication, mRNA transcription and modification in cells. Codon optimization aims to increase the yield of protein expression in different organisms by rebalancing usage of synonymous codons. High quality protein has a wide range of applications, including study of protein structure and function, biological research, clinical medicine.

Higher protein yield in turn can be a valuable feature in a wide range of applications, including…

Codon usage bias

There are 64 codons in total, 61 codons encode 20 amino acids and the remaining 3 are stop codons. Redundant codons exist for most amino acids; multiple different codons encode the same amino acid. In general, the expression of heterologous proteins is negatively correlated with the occurrences of rare codons. In simpler terms, when the codon usage of a target protein in its native organism differs significantly from the average codon usage of the expression host, this could cause problems during expression. Therefore, an effective solution is to replace rare codons in the original sequences with major codons that are preferred in the host.

The secondary structure of mRNA

Translation refers to the conversion of nucleic acid sequences into amino acid sequences, and is greatly affected by complex secondary structures in mRNA. The translation of codons in the non-structural part of the α-helix is the rate-determing step in the translational process. Identifying the hairpin structure in an mRNA can lead to significant optimizations of translational efficiency for that mRNA.

Changing regulatory elements and restriction sites

The process of optimizing protein expression is affected by many factors: not only basic DNA transcription and translation, but also a series of regulatory elements. By incorporating signal peptides, tags, other regulatory elements, or appropriate restriction sites during gene synthesis, expression of synthetic genes in heterologous host organisms can be significantly improved.

Synbio Tech NG^TMCodon Optimization software can intelligently optimize codon usage to effectively improve protein expression, in order to better meet the needs of scientific research and industrial production.

Synbio Technologies’ NG^TMCodon Optimization Software

Synbio Technologies’ proprietary NG^TMCodon optimization software provides customers with free codon optimization services bundled with gene synthesis. Our NG^TM Codon optimization software optimizes gene sequences based on different expression systems to maximize protein expression.

NG^TMCodon optimization software advantages：

Elimination of codon usage bias:

Codon usage bias refers to the differences in frequency of synonymous codons in coding DNA, and exists in a wide variety of organisms. The most frequently used codon is called the optimal codon, and lower frequency codons are called rare codons. Synbio Technologies’ proprietary codon optimization software, NG^TMCodon optimizes the usage of both the optimal codon and the rare codons to achieve superior protein expression.

Fine-tuning secondary structure of mRNA

The secondary structure of mRNAs play an active role in the translation process. The complexity and stability of mRNA secondary structure both greatly affect the smoothness of the translation process, in particular, secondary structure near the ribosome binding site (RBS) is crucial. NG^TMCodon can effectively identify and minimize hairpin structures in mRNA, effectively optimize the original sequence, and generate significant improvements in protein expression.

Removal of negative cis-acting elements and restriction sites

NG^TMCodon was designed to eliminate multiple obstacles during transcription and translation by minimizing negative cis-acting elements that interfere with transcription and translation.

Case study：Targeting protein was reproducibly expressed in E. coli after codon optimization by NG^TMCodon.

NG^TMCodon improves codon usage efficiency

Four Steps of Subcloning Technology Protocol

Subcloning refers to the technique of re-cloning a DNA fragment from one vector to another, so that we can more easily perform analysis, transformation, and recombination of the target gene(s). Subcloning is an important tool in any molecular biologist’s toolkit, helping to elucidate the function of a target gene and to easily analyze its phenotype. There are four steps in the subcloning process: obtain the target fragment, connect enzyme vector and target fragment, transform in host cell, identify and screen.

Subcloning Technology Protocol- obtain the target fragment

Find the target fragment in a gene library
The cDNA sequence was reverse transcribed with mRNA as a template through PCR, so we can obtain the target fragment from the cDNA library
Cut the DNA into many fragments with restriction enzymes and introduce into cells; we can screen for cells containing the target gene(s)
Synthesize the gene sequences in vitro

Subcloning Technology Protocol- connect enzyme vector and target fragment

Choose an appropriate restriction enzyme to cleave the target fragment from the vector. Cleavage in this way usually generates symmetrical cohesive ends, non-symmetric cohesive ends, or blunt ends. For subcloning, non-symmetric cohesive ends are preferred.

Subcloning Technology Protocol- transform into host cell

The two most common methods of transforming the target fragment into cells are transformation / transfection and transduction. In the first method, a recombinant plasmid or phage is simply transformed into a treated host cell; in the second, a host cell is transducted with a virus harboring exogenous DNA. In general, transduction is more efficient than transformation.

Subcloning Technology Protocol-identify & screen

Vectors with recognizable genetic markers can be used to help distinguish and separate cells transformed with recombinant DNA. For example, color can be used as such a marker, as the color of a colony may change in some vectors with exogenous genes. Other effective and widely used screening methods also exist, such as screening with drugs or with selective media.

With our Syno^®2.0 gene synthesis platform and experienced technical team, Synbio Technologies provides one-stop subcloning services including target gene synthesis, vector construction and transformation, identification, and screening.

Synbio Technologies’ PCR cloning and Subcloning Technology

PCR cloning and subcloning technology, first developed in the 1970s, is now a staple in every molecular biology lab in the world. Cloning allows researchers to much more easily understand gene function at a deeper level, and greatly facilitates gene editing. PCR cloning and subcloning technology is not only revolutionary for the field of biology, but has profound implications on fields like agriculture, industry, and medicine as well.

PCR cloning technology

PCR cloning technology is similar to natural DNA replication, and contains three basic reaction steps: modification-annealing-extension. We can obtain a desired target gene sequence with appropriate primer design. PCR can then be used to amplify this gene sequence, preparing it for use in cloning.

Subcloning technology

In molecular cloning, target DNA is assembled into a vector plasmid through restriction enzymes and screening. In subcloning, a gene of interest is transferred from one vector to another. Both processes consist of several key steps, such as screening of the target fragment, cloning vector preparation, transformation/transduction of the product into cells, and screening for cells containing recombinant plasmids.

Both PCR cloning and subcloning technology can insert a target gene into a plasmid of choice in vitro through recombinant technology. The main forms of target gene transfer into a plasmid are transformation and transduction. This allows researchers to have an enormous amount of customization available to them when trying to study a gene of interest, making cloning and sub-cloning two extremely powerful tools in a molecular biologist’s arsenal.

With our proprietary Syno^®2.0 gene synthesis platform, Synbio Technologies can provide one-step services for gene synthesis, vector construction, PCR cloning, and subcloning. Customers only need to offer the sequence information, and we can help design amplification primers and clone the PCR products to the specific sites of the new plasmid. We also provide sequencing services in order to confirm that the correct product was accurately synthesized.

A Brief Introduction of Molecular Biology Techniques

Molecular biology studies the morphology, structural features, biological relevance and regulatory mechanism of biological macromolecules including nucleic acid, protein and others. Molecular biology truly uncovered the mysteries of the biological world from the molecular level; it signaled the transition from passive adaption from nature to actively restructuring and reengineering the nature.

The main research contents of molecular biology span a broad spectrum: applications of DNA recombinant technology (also known as genetic engineering) in genetic engineered drugs; transgenic plants and somatic cell nuclear transfer; gene expression and regulation; structure-function relationship of biological macromolecules; functional genomics and bioinformatics studies.

Molecular biology is the cornerstone of current life sciences and medicine-related disciplines. Molecular biology techniques are the most commonly used tools in life science, however the viability and reliability of experiment result are dependence of many factors, including experience and expertise.

Empowered by Syno^®1.0 and Syno^®2.0 platforms, Synbio Technologies provides comprehensive and accurate molecular biology service for worldwide customers, ranging from DNA cloning, site-directed mutagenesis, vector construction, protein expression, gene/genome sequencing, gene synthesis technology and many other services.

The Wide Applications of Molecular Biology

Molecular biology clarifies regularities of cell and receals the essence of life.Molecular biology technologies have been widely adapted for recombinant protein production, genetic modification of organisms, gene therapy, environmental protection, etc.

Production of recombinant proteins

Recombinant Insulin production

For very long time, the insulin that was used to treat diabetic patients is solely purified from bovine or porcine pancreas. 100kg pancreas can only extract 4-5g of insulin. The development in the field of genetic engineering introduced chemically synthesized insulin cDNA in E.coli and allowed insulin production in microorganisms, yielding 100g insulin from every 2000L microorganism’s culture. The massive industrial scale production of insulin not only solves the yield problem but also drives its price down by 30% -50%.

Genetic engineering drugs

Genetic engineering is used to mass-produce interferon, artificial blood, interleukin, hepatitis B vaccine and many other drugs which played a huge role of lifting the human suffering, improving human health.Genetic engineering drugs has greatly been embraced by man to improve on his well-being.

Genetically modified animal

For certain protein drugs that require complex modifications or are needed in large supply, production in transgenic animals seems most efficient. The current strategy to achieve these objectives is to couple the DNA for the protein drug with a DNA signal directing production in the mammary gland.

Herbicide resistant crops

Herbicide resistant crops are genetically modified to tolerate broad-spectrum herbicides, which kill the surrounding flora, but leave the cultivated crop intact.

Genetically modified microorganism

Microorganisms are most commonly used in genetic engineering due to their inexpensive nature. Cheese production requires the use of a protein called chymosin which is a proteolitic enzyme usually obtained from calf stomachs. Production of chymosin in genetically engineered microorganism provides an alternative way of producing cheese that does not require the sacrifice of large amount of animals. Moreover, microorganism production of recombinant chymosin offers an easy way of increasing the production of chymosin compared to the amount that can be obtained from young calves.

Gene therapy

Gene therapy is a technique to treat genetic diseases by introducing foreign DNA which usually contains a functioning gene to correct the effects of a disease-causing mutation.

Environmental protection

The genetically engineered organisms can be used as bioindicators to readily reflecting pollution level on a habitat, community, or ecosystem. Moreover, these bioindicators are engineered to resist pollutant-leaded mortality and potentially has the capability in bioremediation of toxic chemicals.