Bovine Methane Emission Control

Bovine Methane Emission Control

Part 1

The Proposal

The proposal presented by Dr. Kenmore’s about “cow of future” is explained expertly capturing the conceptual framework of how the project will be aimed at reducing methane emission. Dr. Kenmore has experience in microbiology, and he has cloned Methylococcus capsulatus and then expressed the genes to Escherichia coli. Moreover, he has an interest in probiotics since he has been consuming on kefir and yogurt, which makes him passionate and willing to bring a revolution to control methane gas that has contributed to global warming. In his project, he has tried to express critical concepts that need to be put in place to contribute to reducing global warming as a result of methane emission from bovine. Global warming has been one of the major concerns of global issues that have lead to climatic change and health issues.

Part 11

Underlying Science

Question 1: The arrangement or sequence of pmoC, pmoA, and pmoB genes DNA? The genes contain three-gene operon cluster pmoC, pmoA, and pmoB proteins. This operon code for specific integral membrane polypeptide that is used to form (particulate methane monooxygenase) MMO the genes occur in clusters. The pmoC contains six membrane rotating regions and no classical N terminals sequences. Besides, pmoA has upstream region 3’ that has an N terminal that provides methionine amino acid (Henard et al. 2016).  And then, the pmoB gene is found downstream and diverges at the N and C terminal of protein codon.  The genes for pmoA and pmoB are identical and are encoded by one gene. The pmoCAB clusters and pmoC genes form a bath that clones for phage of Escherichia coli.

b.) Which would you select as the most accurate and useful depiction of the sequence of the pmoC, pmoA, and pmoB genes in the M. capsulatus chromosome?

The answer is diagram D that is drawn in triad arrangement, and this makes it easy to identify the locations of various genes and direction of translation and transcription. Different direction has been shown to represent genes coding for different functions like respiration, taxis, and metabolism. The diagram shows the actual appearance in the genetic representation of linear-direction from pmoC, pmoA, and pmoB, which are easy to explain and understand the process of transcription and translation.

Question 2: The gene that includes several necessary sequences that ensure translation and transcription of the gene: promoter, termination and initiation courses start and stop codons, RBS, what is the role of each of these?

• Promoter sequences are the portion of a DNA initiates and begin transcription of specific genes. The promoter sequences are located close to transcription sites of codons and are located upstream and on the same DNA strand.
• The initiation sequence begins the transition of single stranding RNA to double-stranded DNA. The polymerase of the RNA acts as a promoter of the series hence promoting transcription and translation process.
• Termination sequence is a codon of RNA that marks the end of the transcription process of genetic information.
• Start and stop codons the start codon indicates the site in which translation on a sequence of protein will begin and replicate analogous genetic codes. While the stop codon, it marks the point where the protein sequences will end during translation to promote base pairing.
• RBS sequence is used in the initiation phase of transcription, where it recruits the start codon. It comprises nucleotides that are placed upstream to start codon of the messenger RNA and then feeds it with ribosomes that are the replicating tool during transcription.

Question 3: The essential sequences from Question 2 above located? Amino acid 1557

54. How many promoters are necessary to propagate transcription of the operon pmoCAB? One promoter molecule in the transcription phase is required, and that is P54.
55. How many molecules of mRNA will be available when this operon is transcribed?

One molecule would be produced after transcription.

1. The number of ribosome binding sites that will be available on the RNA produced during transcription of the PMO operon?

Three binding sites would be present.

1. Indicate the numbers of stop codons and start codons that will be available on the RNA produced during transcription of the PMO operon?

There will be three stop codons, and three start codons.

1. Which letter indicates a sequence of DNA where RNA polymerase binds to ensure transcription of pmoA, pmoB, and pmoC as a unit?

(b, c, g, h)

1. Which letter represents the initiation transcription site? (a, b, e, f)
2. What representation is transcription termination? ( d, e, f)
3. Find start site sequences encoding of translation? (c, h, I, j) What would be the location(s) of stop codons are DNA sequences encoding translational? ( K, l, m)
4. Which letter in the diagram indicates the 5′ end of a DNA strand?

The end of a strand of DNA is marked by 5′ on the left, and 3′ is on the right. Moreover, it can also be 5′ and the letter n, which indicates an end. In the diagram, it is a letter (a)

Part 111

A mistake

Question 4: Mistake identification of the nucleotides in the gene was changed, as indicated in the mistake sequence? Silent mutation Explain:  

These are silent mutations since the arrangement starts from query compared to the subject sequence where there is no much difference (Strong et al. 2016). The site at subject and question ends with A, thus no difference or change in silent mutation.

1. The sequenced DNA shown indicates the mRNA sequence be when the template DNA is transcribed? (First 60 bases given). The amino acid sequence when the mRNA is translated? (first 15 amino acids given).
2. AUG CGU CCG UCA ACC CCG ACU GUU CCG CCA AGA ACU CCG GCC UUC UGU GGC GGG AUG ACG UUG AGC ACA UCG GCA CAG AAA GUG UCG AAC UCG GUU UCG ACG GUU CGC CCG GCG GCA UCG AUC CGC UCC AGC CGU CCC UCU UUU UCC GAG GAA
3. GUCGACUGGG CACCAGCCGG AUGCGUCCGU CAACCCCGAC UGUUCCGCCA AGAACUCCGG

2. Data analysis of how the protein products of this DNA sequence differ from the PmoC that would be made if the gene did not have a mutation?

GTCGACTGGG CACCAGCCGG ATGCGTCCGT CAACCCCGAC TGTTCCGCCA AGAACTCCGG

1. Guess as to the functionality of this altered protein associated with PmoC.

The prediction indicates that in silent mutation, even if there is a slight change in amino acid bases, no change in functionality, so nothing is affected because the amino acid retains the function.

Part IV: The process of cloning

1. The cloning vector analysis of plasmid vector: pCK1 coli. What is the role of each labeled section? The relevance of the cloning experiment?
2. Ligation is a process where there is a linkage of two linear DNA strands resulting in one piece, and the process is catalyzed by DNA ligase. M. capsulatus chromosome is transformed then so that it can be used as phage in E. coli, which expresses the desired genes.
3. Amplification of PCR is a process where the tubes containing the M. capsulatus are placed in a centrifuge or PCR to increase the potential of ligation and express the genes.
4. Digestion restriction also is a process whereby the DNA strand is cut to introduce a new strand, which is used as phage. Digestion allows a particular gene to be translated during the transformation process.
5. The transformation involves changing the genetic pattern of the M. capsulatus by the introduction of a new linear DNA strand that is used to express the desired trait in a phage.
6. Antibiotic resistance: is observed when the growth media containing the LB and ampicillin is used to culture from the tubes that have E. Coli, which is used to expressed genes of a phage.

1. Cloning Review process.
2. Which step in the diagram indicated, would DNA ligase be applicable? It happens in the 1^st
3. Which step in the diagram would an endonuclease restriction be used? It is done in steps two and three.
4. The ampicillin resistance gene is significant in which step? Step five is the best stage to find ampicillin resistance.
5. The transformation representation of bacterial vegetation is indicated in which step? Step 4 denotes transformation.
6. What is added to the growth media to enable the pmoCAB operon expression? The growth that ensures phage expression should be fortified with LB and ampicillin

2. The genetic transfer can occur naturally between bacteria by three various mechanisms. Explain the arrangement they would occur

First, it would occur through a transformation where LacZ is ligated from the genes of the M. capsulatus, and then another desirable gene is attached in the space where ligation took place (Henard et al. 2016). Subsequently, transduction follows where bacteriophage will be instituted in E.coli to express the proper traits to digest cellulose and prevent methane emission and release.

Reference

Henard, C. A., Smith, H., Dowe, N., Kalyuzhnaya, M. G., Pienkos, P. T., & Guarnieri, M. T. (2016). Bioconversion of methane to lactate by an obligate methanotrophic bacterium. Scientific reports, 6, 21585.

Strong, P. J., Kalyuzhnaya, M., Silverman, J., & Clarke, W. P. (2016). A methanotroph-based biorefinery: potential scenarios for generating multiple products from a single fermentation. Bioresource technology, 215, 314-323.