(b) explain how CRISPR-Cas9 can be used to edit genomes and potentially cure dis
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Question
(b) explain how CRISPR-Cas9 can be used to edit genomes and potentially cure diseases(c)describes how Nucleic Acid Hybridization can be used to diagnose a patient who has syphilis ( this bacterium can not be grown on agar)
(d) identify the similarities between DNA replication and Polymerase chain reaction.In your own opinion, without the knowledge of DNA replication would scientists have been able to develop PCR? Explain your reasoning In the figure below identify 6 ways scientist have applied the knowle that we have discussed so far about microorganisms (bacteria) or molecular biology dge of processes or properties Example: The fact that bacteria can grow to high numbers in liquid culture allows scientists to grow large numbers of bacteria that produce (secrete) a desired chemical as a step to mass produce the chemical economically (for less money) enzymes3 human isulin protein & insulin ready for
Explanation / Answer
1a) Required ways from the figure given:
(i) Polymerase chain reaction: To produce a number of copies of required DNA sequence.
(ii) Recombinant DNA Technology: A process of combining required gene at a specific site in already occuring genome.
(iii) Transformation : This is a process by which plasmid is introduced into a bacterial cell.
(iv) Action of various enzymes like ligases or endonucleases: Restrication enzymes like Endonucleases specifically those which cleave genetic material at a specific site for eg. EcoR-I etc and Ligases (those which catalyze the rejoining of genetic material)
(v) Expression of plasmid genome by bacteria: The bacterial cells when have the plasmid, express the genes on it along with its own genome
(vi) Fermenatation: In this the genetically engineered bacteria are provided with nutrient media and allowed to multiply under anaerobic conditions.
1b) CRISPR-Cas9 is an enzyme which aids in changing the genome, causing the formation of different proteins in the organism and hence modifying the response to different conditions. The basic origin of its importance was based on the provision of immunity to bacteria against different bacteriophages. The modifications in genome caused by this enzyme help the bacteria to remember the process of action by which it can then get rid of the viral genome when infected by the same. This technique can be utilized to edit the genomes of other organisms and hence can cure a number of diseases.
Methodology: CRISPR-Cas9 works in association with specific RNA sequence which is known as guide RNA's, whose function is to change the base pairs of a given mRNA or DNA on the basis of complementary Watson Crick base pairing rules. Cas9 dependent RNA sequences which can bind the enzyme can be created in the laboratories. The RNA should be such that they are capable of binding to specific DNA sequence of the target organism/pathogen. Once the Cas9 systems gets attached to the site, then it causes the unwinding of DNA from that site followed by binding of guide RNA at that site and then causing the changes in DNA of the pathogen by the mechanism taking place in repair at the time of DNA replication. Once the genome gets changed, it may bring about different results like it may prevent normal replication in the pathogen or it may prevent pathogen to translate certain sequence and so preventing the release of toxin responsible for pathogenic action. In this way, this system can be used for curing the diseases.
1(c) Nucleic Acid hybridization is based on complementary base pairing binding between the different DNA or DNA-RNA sequences. This technique can be used in diagnosis of different diseases.
In case of syphilis, which is caused by Treponema pallidum, the pathogen is uncluturable on agar plates and hence its source has to be maintained by keeping it in the host For that Rabbits are infected with syphilis and are used to collect the sample of pathogen. The rabbits are chosen because (i) they get infected by same species as humans (ii) The infection in rabbits can be easily cured.
By taking the genetic material of the lab grown pathogen and from that of the infected organism (whose diagnosis is to be done) are allowed to hybridize on inert support and unhydridized/unbound material is removed. If the similar genome is detected, the patient is daignosed with syphilis.
This is done in following way: The genetic material
1(d) Similarities between PCR and DNA Replication:
(i) both the process should have an initial small sequence called the template
(ii) Both follow same base pairing rules.
(iii) Addition of nucleotides in both the processes takes place in same direction i.e. 5'-3'.
(iv) Both require DNA polymerase but the unwinding and ligation in PCR is enzyme independent
Knowledge of DNA replication was mandatory for scientists to develop PCR because PCR is actually DNA replication but of a specific gene. The processes involved in PCR like Initiation, Unwinding, Annealing etc are similar to those found in DNA replication process. So, without the knowledge of various replication steps like formation of primer, its binding and then base pairing direction, it was not possible to develop a technique like PCR.
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