CRISPR-Cas9 and Recombinant DNA in Genetic Engineering

What is recombinant DNA (rDNA)?

Recombinant DNA (rDNA) is produced by combining genetic material from two or more different sources.

What is the purpose of recombinant DNA?

rDNA allows a gene from one organism to be moved into, and expressed in, a different organism.

What are two important tools used to create recombinant DNA?

Endonucleases (such as restriction enzymes or the CRISPR-Cas9 system) and DNA ligase.

How does CRISPR-Cas9 function in bacteria?

CRISPR-Cas9 acts as a primitive adaptive immune system that remembers past viral infections.

What is a bacteriophage?

A bacteriophage is a virus that infects and replicates within bacteria and archaea.

What is a protospacer in the context of CRISPR?

A protospacer is a small segment of viral DNA that is cut out and inserted into the bacterial chromosome.

What is the CRISPR array?

The CRISPR array is part of the bacterial chromosome that contains spacers and identical repeats.

What is the role of RNA polymerase in CRISPR?

RNA polymerase transcribes the CRISPR array to produce pre-crRNA, which contains both spacers and repeats.

What is single guide RNA (sgRNA)?

sgRNA is a synthetic RNA molecule designed to guide Cas9 to a specific DNA sequence for cutting.

What does Cas9 recognize in a DNA sequence?

Cas9 identifies a PAM (protospacer adjacent motif) region in the DNA sequence.

What happens when Cas9 cuts DNA?

Cas9 causes a double strand break in the DNA if the sequence upstream of the PAM site is complementary to the guide RNA.

What are the two ways a cell can repair a double strand break?

Non-homologous end joining and homology directed repair.

What occurs during non-homologous end joining?

Enzymes hold the broken ends together, and ligase joins the two ends, which can lead to mistakes.

What is homology directed repair?

A repair method that uses the complete DNA sequence on the homologous chromosome to rebuild the broken strand with fewer errors.

What are some applications of CRISPR-Cas9 technology?

Applications include gene knockout, introducing specific mutations, editing faulty alleles, and modifying crops.

How can CRISPR-Cas9 advance animal welfare?

By creating traits such as hornless dairy cows.

What is the significance of CRISPR-Cas9 in agriculture?

It can be used to modify crops to increase their nutritional value.

What is the role of tracrRNA in CRISPR?

tracrRNA binds to the repeat sections of pre-crRNA to help form the functional gRNA.

What happens if the DNA sequence is not complementary to the gRNA?

Cas9 will not cut the DNA and will move on to locate another PAM site.

What is a spacer in the CRISPR system?

A spacer is the viral DNA segment that has been integrated into the bacterial chromosome.

What is the potential impact of CRISPR-Cas9 on inherited diseases?

It can be used to edit faulty alleles in individuals with severe inherited diseases.

What is the purpose of creating mutations using CRISPR-Cas9?

To disable gene function or produce a STOP signal in a gene.