Genotyping, Pharmacogenomics, and Antimicrobial Peptides Analysis Assignment Sample

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1. Introduction

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Section A

a.

The genotype of CYP2D6 can be possible with the help of the genotyping process. This genotype identification was possible from the post mortem report of the 6 years old child. The evaluation of the genotype is also related to the ultra-rapid metabolizer phenotypes. People who are ultra-rapid metabolizer phenotypes have such a higher enzymatic activity rating than 2.5, which is frequently attributable to an enhanced nucleotide sequence of the CYP2D6 gene. The phenotype is thought to just be exhibited around 1% to 2% of people, but the incidence varies amongst populations. By the use of the AmpliChip CYP450 Test, the identification of the genotype can be possible. The genotyping method is possible by the identification of the genomic DNA (Gusella et al. 2020). The main protocol for the identification of the genotype is PCR. With the help of the polymerase chain reaction, the amplification of the DNA can be possible that is helpful for the sequencing of the DNA.

Figure: Protocol for identification of the CYP2D6 genotype

(Source: https://www.researchgate.net)

For the identification of the CYP2D6, the protocol is mentioned in the above diagram. In genomics, the identification of the particular sequence of the gene such as CYP2D6 can be possible by the analysis of the particular technique that is known as RFLP. in this technique the desired gene can be identified by the use of the hybridization technique. From this diagram, it can be explained that the identification of the genomic DNA is the first step. After the identification of the DNA sequence from the whole DNA the desired sequence can be isolated by the use of the restriction endonuclease. The steps in these diagrams are under the RFLP technique. After the identification of the DNA, the PCR is carried out for the amplification of the DNA and this PCR technique is also required to be carried out by the specified time. For each of the cycles of the PCR technique, the generation of the two double-stranded DNA can be possible. In this way according to the requirement of the amount of the DNA strands the numbers of the cycle for the PCR technique are required to be continued (Okat et al. 2018). The gene sequence that is related to the CYP2D6 can be identif\ied and the amplification of the sequence is important. After that, the identification of the presence of the specific gene can be possible by the hybridization technique. After the hybridization technique, the transfer of the desired sequence of the gene can be detected.

b.

The correlation of the mentioned phenotype can be mentioned below. For the identification of the genotype, the sequencing of the DNA sample is important. By the arrangements of the gene sequence, the type can be identified and identification of the genotype can be possible. By this table, the correlation of the metabolism and the medication dosage can be presented below.

c.

The drugs are applied to 6 years-old children for the treatment of the postcondition of appendicectomy. For this treatment, the liquid codeine with its proper dosage according to the age of the child is applied. It is one kind of syrup and in this medicine, ibuprofen, and aspirin can be present. In the post-mortem, the presence of Morphine is also detected. This Morphine is one kind of drug that is used to relieve the pain that can be present within the child after the surgery for the removal of the appendix. By the application of the medicines, the safety of the drug can be improved and the new target of the drug can also be evaluated. Morphine is the drug and the target site of this drug is the nervous system the prevention of the pain can be possible by the activation of the drug to the target side that is the nervous system. In the case of the presence of side effects the use of the drugs is required to be prevented (Asaduzzaman, 2018). The dosage of the medicines is also important to be maintained according to the age of the child. The dosage can vary in effectiveness as per the route of the administration.

For the personalization of the medicines, pharmacogenomics can be effective. According to the individual genotypes, the tailoring of the drug therapy can be possible with the help of pharmacogenomics. The development of a new drug can be possible, which is very important for the presence of drug resistance within the people. The Discovery of new drugs and the development of the process of metabolism can also be possible with the help of pharmacogenomics. Within precision medicines, pharmacogenomics plays an important role. Pharmaceutical companies can also be properly guided by these techniques for effective drug development and also for the new type of drug discovery. With the help of pharmacogenomics, the expression of the proteins and other enzymes within the person can be identified. This affects the metabolism of the drug can also be identified and the solution can also be possible. With the help of pharmacogenomics, the identification of the specific drug can be possible with the help of the physician and this identification can be done based on the genetic makeup of the patient. Prescription of the right dose of the drugs can be possible by this technique and due to this the efficiency of the drugs can be increased (Hassan et al. 2021). The avoidance of ADR of the patient can be possible with the help of this technique. This pharmacogenomics is one kind of study through which the analysis of the genomic sequence of the patient can be possible. From this analysis the4 response of the patient to the medications can also be evaluated.

3.

a. i.

The development of antimicrobial peptide-based drugs that are also known as AMPs is effective for the host defense mechanism. For the improvement of the mechanism, the research of the scientists is developed for making AMPs. The depreciation of the novel peptide can be A3 and that is effective for the treatment of drug-registrant infections. The derivation of the new components can be possible by the deduction or the substitution of the amino acids. The changes of the distribution and also the composition of the components of the amino acids sequence within the polypeptides the effect of the peptides can be changed. In the development of the AMPs, the production of the new antibiotics can be possible and for this,

Some of the challenges can be faced by the researchers. The technical challenges for the development of the AMPs can be mentioned. The problem can be incorporated into the formation process of the peptides. Generally, the process of fermentation is used for the formation of the AMPS. In this case, the organisms can be used. This fermentation can be carried out within large-size containers. In this case, the control for the temperature, concentration of the oxygen for the production of the organisms, and maintenance of the nutrition label and pH is also effective (Tang et al. 2021). Different kinds of technical barriers can be the changes in the activity of the enzymes within the organisms. With this, the modification of the target and the membrane barrier can be present that can be the major challenges for the development of the peptides. By taking the longer dosage of the antibiotics the immune system of the body can also be affected. In case of the changes in the mechanisms of growing bacteria the development of the antibiotic can also be the main challenge. The toxicity of the antibiotic is also required to be checked otherwise the side effects rate within humans can be increased.
Some of the technical challenges can be the formation of antimicrobial peptides with the help of the development in nanotechnology. Inclusion of the AMPs within the drug delivery system is also challenging and it is responsible for the low amount of efficiency within the encapsulation. Due to the presence of the structural complexity within the AMPs, the delivery system of the AMPs within the drug can be challenging. The main technological problem is the presence of high-cost that is required for the transfer of the AMPs within the drug delivery system. After the transfer of the AMPs, this can be inactivated by the enzymatic reaction within the drug and for this, the high rate of instability of the AMPs can be a major disadvantage or the technical challenges within the drug delivery system (Biswaro et al. 2018). Some of the technical problems can be present within the experiment of the in-vivo system. For this, the development of the effective result cannot be possible that can be mentioned as the effective problem for the development of the AMPs for the clinical development. With the help of antimicrobial peptides, the prevention of the microbial effect can be possible.

a. ii.

Some of the financial challenges for the development of the AMPs can be the high-level cost of the medicines. For this, the rate of the sales of these medicines can be lowered and the rate of profitability can also be reduced. The development of antibiotics or antimicrobial peptides is a time-consuming process and for this, the expense for the production of the antibiotics can be increased. But in the case of the necessity of the use of antibiotics reasonable prices are required to be maintained so that the people can be benefitted from the use of these antibiotics. Due to the increased cost of antibiotics, the health services cost can also be increased. The initiatives of the international level can be effective for bearing the production cost of antimicrobial peptides. Due to the high rate pricing, the availability of antimicrobial peptides can be affected. The shortage of medicines in the market is also possible. These can be stated as the effective problems or challenges in restricting access to the peptides (Chen, and Lu, 2020). For this, the treatment of the common infections that are caused by the bacteria can not be treated properly. In the case of the resistance of the bacteria in the present antibiotic the development of the antibiotics is important but, in this case, the analysis of the mechanism of the bacteria is required to be investigated. For providing more time the expense rate is increased. The financial challenges are also borne by the patient. Due to high pricing, the economic condition of the patient is also affected. In the case of the low rate of acceptance, loss of the economy of the production company is also the major challenge.

The financial challenges for the evaluation of the clinical development of the AMPs can be mentioned as this process is time-consuming. Despite the success of this technique, it cannot be possible. In case of the instability of the AMPs within drugs, the procedures can be mentioned as the failure of the transfer of AMPs. The financial challenge is the main problem. For any kind of experiment, financial investment is necessary. For the use of the AMPs within nanotechnology the financial investments are required to be high. In this case, the successful result is important for financial success also. The development of the clinical implication of AMPs can be possible by the expense of the mines and also it is a time taking process (Salas-Ambrosio et al. 2020). For this, the financial challenges can be effective for the development of the effective nature of AMPs and the clinical development of AMPs. For clinical development, the analysis of the peptide sequence is also important and in this way, the selection of the peptide sequence and its transfer to the drug delivery system can be effective.

b.

In the function of the antibiotics or the antimicrobial peptides, the generation of the problems can be incorporated. The smart and flexible regulation of antibiotics is effective for the prevention of bacterial infection. The mechanisms of the action of the drug are required to be effective. So that the doctor can prescribe the newly formed drug to the patient for its effectiveness to the target side of the patient. The changes in the mechanism of the action of the organisms can be the major problem for the generation of the resistant action of the bacteria. Due to the resistance problem, the staying period in the hospital by the patient can be incensed which can lead to the economic and financial problems of the patient family (Mahlapuu et al. 2020). The rate of mortality within the patient can also be increased, which can be referred to as the major regulatory problem.

Some of the regulatory challenges can be mentioned. The presence of a low level of awareness in the case of self-medication can be the regulatory challenge of the clinical development of AMPs. The lack of a national-level surveillance system can be another challenge for the clinical development of AMPs. The absence within the legal boundaries, due to the lack of the financial resources for the development of the clinical experiment the challenges can appear (Wang et al. 2019). The presence of the resistant data of the microbial infections is important. Due to the lack of the proper resistant data the development of the AMPs can be affected. The lack of waste and sanitation management can be another challenge for the clinical development of AMPs.

Figure 2: Technical, financial, and regulatory challenges for the development of antimicrobial peptides

(Source: https://www.mdpi.net)

4.

a. i.

Genome sequencing is an effective tool that is important for the isolation of the genomic DNA and also effective for the evaluation of the pathogenesis of the organisms. The sequencing can be accurate with the help of the Sanger sequencing methods. This is one kind of automated method of sequencing. The bands of the DNA sample that are moved through the gel can be read through the technique based on computer reading. The terminal dDNTPs can be identified with the help of the fluorescence tag. A laser activates the fluorescence tags within every band, and often a machine analyses the light produced. The nested fragment of the DNA can be generated according to the sequence of the template DNA in this sequencing method (Cheng et al. 2021). The sequencing of the template DNA fragment can be possible by the replication process. At the time of the replication, interruption is required after four bases for the completion of the sequencing method.

Figure 3: Sanger method of Genome sequencing

(Source: https://www.onlinebiologynotes.net)

Each of the steps for Sanger sequencing can be mentioned as three steps. The denaturation of the double-stranded DNA. Due to this denaturation, the formation of the single-stranded DNA can be possible. The corresponding primer that is designed according to the template sequence of the single-stranded DNA can be attached to the end portion of the sequence. The dNTPs solution can be taken and from this, the particular ddNTPs can be added for the PCR reaction. The separation of the DNA can be possible with the help of the gel electrophoresis technique and from this, the detection of the sequence can be possible. The read capacity of this Sanger sequencing method is about 800 base pairs (Schmid et al. 2022). Within these base pairs, about 500 to 600 base pairs are contained with non-enriched DNA. The length of the Sanger sequencing is the advantageous part by which the repetition that can be present within the genome sequence can be identified. The Sanger sequencing result is effective and restricts the size of the genome to 10 kb.

a. ii.

The principle of next-generation sequencing can be abbreviated as the NGS sequencing method is somewhat similar to the Sanger sequencing method. The principle of this technique is based on the capillary action of the electrophoresis technique. Whenever the segments of the genome strands are spliced to a target sequence, the nucleotides in each segment are recognized by transmitted signals. Four processes can be possible within this technique and the first step is the preparation of the library, the second step is the cluster generation, the third step is the sequencing and the fourth step is the formation of the alignment and the analysis of the revealed data. This technique focuses upon sequencing-by-synthesis (SBS) with changeable colorants, which allow individual bases to be identified as they can be inserted into the DNA sequence of the strands (Cervantes et al. 2021). Purified genomic DNA is ultrasonically fragmented into DNA fragments ranging from 200 to 500bp. The main methods for the NGS workflow can be mentioned. These methods can be the preparation of the library of the genome, the sequencing of the genome by which the sequence analysis of the gene can be possible. The third process can be the analysis of the data that is obtained from the sequencing method. With the help of these processes, the idea for the formation of the set for the NGS workflow can be possible. The formation of the cluster generation can also be possible by this technique (Tenedini et al. 2022). The validation of the NGS workflow can be possible by defining the numbers of the sample, repeatability, and reproducibility of the detection of the variants within the genome, by the set of the reference range, clinical specificity, and sensitivity, establishment of the PPV, and PPA. by this way the analysis of the sequencing and the next-generation sequencing can be possible.

b.

With the help of the NGS technique, the sequence of the DNA can be possible for the organisms that are responsible for causing the infection. The information about the run of the single sequence can also be possible. From the data about the detailed sequence of the DNA of the organisms, the pathogenesis factor and the virulence factor of the organism can also be understood. This information is effective for the understanding of the types of infections that are created by the organisms. By understanding the virulence factor of the organism, the development of the antibiotic against the infection of the organisms can be possible. The expense is for sequence, information transport, analytics should keep falling, and DNA purification. Especially, library preparations for sequencing analysis would become even faster and more accurate. Current lengthy technology's high level of failure levels, on either hand, will keep improving until it becomes as precise as brief technologies. In this technology the time both will become redundant. Lengthy innovations will routinely close genome sequences, allowing for compatibility to reference known molecular genotyping methodologies including the locus of the gene variable amount of the tandem repeats of DNA sequence and gel electrophoresis, making sure that history data produced by such methodologies is retained.
With the help of this technique, the sequence of the organisms of the next generation can also be possible and for that, the antidote for the prevention of the organism's infection can be possible with the help of this sequencing method. The preparation of the library of the genome is effective for the use of the primer sequence that is required in the amplification part of the sequencing method. In this step, the preparation can be possible with the help of the ligase and different restriction enzymes that can act at the specific cutting site of the DNA. The sequencing can be possible after the amplification step (Besser et al. 2018). After that, the band that is obtained by the capillary action within the agarose gel electrophoresis can be read with the help of the computer. The sequence of the virulence factor of the organisms can be detected. By this detection, the formation of the complementary sequence can be possible that can be used in the formation of the drug or antibiotic for the prevention of the infection that is caused by the virulence factor of the particular bacteria. In this way, this sequencing method is effective for the prevention of bacterial infection and its outbreak condition also.