@hammouda
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Instead of performing one reaction per well, dPCR involves partitioning the PCR solution into tens of thousands of nano-liter sized droplets, where a separate PCR reaction takes place in each one. The partitioning of the sample allows one to estimate the number of different molecules by assuming that the molecule population follows the Poisson distribution, thus accounting for the possibility of multiple target molecules inhabiting a single droplet.
dPCR measures the actual number of molecules (target DNA) as each molecule is in one droplet, thus making it a discrete “digital” measurement. It provides absolute quantification because dPCR measures the positive fraction of samples, which is the number of droplets that are fluorescing due to proper amplification. This positive fraction accurately indicates the initial amount of template nucleic acid. Similarly, qPCR utilizes fluorescence; however, it measures the intensity of fluorescence at specific times (generally after every amplification cycle) to determine the relative amount of target molecule (DNA), but cannot specify the exact amount without constructing a standard curve using different amounts of a defined standard. It gives the threshold per cycle (CT) and the difference in CT is used to calculate the amount of initial nucleic acid. As such, qPCR is an analog measurement, which may not be as precise due to the extrapolation required to attain a measurement.
dPCR measures the amount of DNA after amplification is complete and then determines the fraction of replicates. This is representative of an endpoint measurement as it requires the observation of the data after the experiment is completed. In contrast, qPCR records the relative fluorescence of the DNA at specific points during the amplification process, which requires stops in the experimental process. This “real-time” aspect of qPCR may theoretically affect results due to the stopping of the experiment. In practice, however, most qPCR thermal cyclers read each sample’s fluorescence very quickly at the end of the annealing/extension step before proceeding to the next melting step, meaning this hypothetical concern is not actually relevant or applicable for the vast majority of researchers. dPCR measures the amplification by measuring the products of end point PCR cycling and is therefore less susceptible to the artifacts arising from impaired amplification efficiencies due to the presence of PCR inhibitors or primer template mismatch.
qPCR is unable to distinguish differences in gene expression or copy number variations that are smaller than twofold. On the other hand, dPCR has a higher precision and has been shown to detect differences of less than 30% in gene expression, distinguish between copy number variations that differ by only 1 copy, and identify alleles that occur at frequencies less than 0.1%.
Source: Digital polymerase chain reaction – Wikipedia
I received this from Medscape medical news:
Genetic Profiles Affect Smokers’ Lung Cancer Risk (medscape.com)
You should not go for the complementary strand.
The DNA coding strand has the same sequence like mRNA except that U is replaced by T.
Please examine the attached file showing a table of genetic code (codons).
Look at your coding DNA strand, starting from 5′-end find the first ATG sequence that represents methionine codon and the first amino acid in sequence. Count the triplets (codons) which follow this starting codon until you reach a stop (termination), which is not counted.
The attached table shows that termination codons are UAA, UAG, and UGA. In the DNA coding strand, these are TAA, TAG, and TGA.
Good luck.
Actually, the axolotl (Ambystoma mexicanum) was previously known for having the largest genome among animals with 32 billion base pairs. However, recent research shows that the Australian lungfish (Neoceratodus forsteri) has replaced the axolotl as holding the record for the “largest genome in the animal kingdom.
The Australian lungfish (Neoceratodus forsteri) has the largest genome of any animal sequenced so far with 43 billion base pairs. This is around 14 times larger than the human genome.
Reference: Giant lungfish genome elucidates the conquest of land by vertebrates
Most probably you are misinterpreting your results. Many of those who go for the direct-to-consumer DNA testing lack basic knowledge about the human genome. Many do not realize that one person’s DNA is less than 0.1%. different from that of another person. Getting their knowledge from the media and science fiction, they think they have got a quite unique DNA. I have encountered several questions led by this belief. It is the responsibility of the providing companies to educate their consumers to minimize such questions.
This answer is from Quora:
Digital twinning is a technology that involves creating a digital replica or “twin” of a real-world system or process, such as a biological or medical system. Digital twinning allows researchers and practitioners to study and analyze the behavior and performance of a system in a virtual environment, using data collected from the real-world system.
The so called “warrior gene” is the R2 variant of MAOA gene. This variant is of low activity, meaning that it produces less of the protein “monoamine oxidase”. This protein is an enzyme that catalyzes inactivation of the neurotransmitters catecholamines and serotonin. Therefore, low activity of the gene is postulated to cause an increase of the levels of dopamine and norepinephrine. Some studies have linked this condition to increased aggression upon provocation.
The pitfall here is declaring aggressors to be not responsible for their action. Lawyers could use this argument in favor of their clients who committed violent crimes. In a 2009 criminal trial in the United States, an argument based on a combination of “warrior gene” and history of child abuse was successfully used to avoid a conviction of first-degree murder and the death penalty.
https://qr.ae/prpiJlThat is right, especially when using lentivirus for delivering the gene of interest to the host cell. Lentivirus is a retrovirus (RNA that is integrated as its complementary DNA in the host genome). Random insertion of the virus in the host genome may cause insertion mutagenesis. An oncogene may be activated and cause cancer. Early trials for treatment of boys with X-linked severe combined immune deficiency showed the development of some leukemias (blood cancers). Therefore, stringent safety studies should be conducted for new gene therapies, and the patients should be monitored for years after the treatment.
By gene therapy, a new functional gene is introduced into the body cells. It produces the protein that was supposed to be produced by an innate gene. Due to the fact that the innate gene is missing, non-functional, or malfunctional, gene therapy becomes a must.
As with any medical procedure, some complications may arise. The body may react adversely to the newly produced protein, considering it a foreign protein. This may lead to failure of gene therapy, with immune reaction of the body against the produced protein.
Complications may arise from the gene-delivering vector. A common type of vector used is adeno-associated virus (AAV). This vector is used to carry the required gene to the target cells. Although the manufacturers ensure that the virus used is infective but not pathogenic, complications still are encountered.
Guidelines should be followed before, during, and after gene therapy. However, this does not guarantee a zero per cent complications. Recently, we have had a shocking news of two fatalities resulting from gene therapy of spinal muscular atrophy:
Death after Zolgensma – Two Cases Acknowledged – Bioinformatics Hub
This $2.1 million gene therapy is the promising life-saving one-time gene therapy for patients with SMA. Thousands of patients have been successfully treated according to the manufacturer, Novartis
The bottom line is: Medicine is not Mathematics.
Fluorescence-activated cell sorting (FACS) is a specialized type of flow cytometry. It provides a method for sorting a heterogeneous mixture of cells into two or more containers, one cell at a time, based on the specific fluorescent characteristics of each cell.
As in ordinary flow cytometry, the cells are passed in a narrow stream of fluid and subjected to a laser beam, one at a time. The stream is then divided into droplets, each containing one cell. The cells are pretreated with fluorescence-tagged antibodies against specific cell surface antigens. A cell with the specific surface antigen will give a specific fluorescence. Once the fluorescence is detected, a charger will give the droplet a positive or a negative charge. The charged droplets are deflected in an electric field towards a specific container. Thus the heterogeneous cell mixture is separated into two or more containers according to the cell surface antigen type.
This technique is useful for research in the fields of cell biology and oncology.
