PLTW Medical Interventions Unit 2.2 Review

Plasmid

- Gene of any size

- No cell specificity

- No immune response

- Does not integrate into the host genome

- Less efficient at entering cells

Liposome

- Gene of any size

- No cell specificity

- No immune response

- Does not integrate into the host genome

- Less efficient at entering cells

Herpes Virus

- Gene up to 20,000bp

- Nervous system cells

- Immune response

- Does not integrate into genome

- Less efficient at entering cells

Adeno-Associated Virus

- Gene up to 5,000bp

- No cell specificity

- No immune response

- Integrate into genome

- Efficient at entering cells

Adeno Virus

- Gene up to 7,500bp

- No cell specificity

- Immune response

- Does not integrate into genome

- Efficient at entering cells

Retro Virus

- Gene up to 8,000bp

- Only diving cells

- Immune response

- Integrates into genome randomly

- Less efficient at entering cells

Lenti Virus

- Gene up to 8,500bp

- No cell specificity

- No immune response

- Integrates into genome randomly

- Efficient at entering cells

Invivo

- Inside patient

- Pros: Benefits occur more rapidly

- Cons: Greater chance of immune response, vector may enter unselected cell, cannot be analyzed to see results

Exvivo

- Outside of patient

- Pros: Less likely to cause immune response, can ensure cell selection and ensure results of therapy

- Cons: Patient has to wait for benefits of therapy

Concerns of Gene Therapy

- Immune response

- Selected cells

- Function of vector

How is the new gene transferred into the hosts cells?

There are two different ways that the DNA can be integrated into the cells. When using a virus as a vector, they will attach to the outside of the cell (as they usually do) but will release the modified DNA that will code for the correct protein in the cell. As for the use of other vectors, plasmids would be taken up and integrated by surrounding cells (through pili)

What does CRISPR stand for?

Clustered Regularly Interspaced Short Palindromic Repeats

Where does CRISPR come from?

It is a system used by bacteria to defend themselves against viruses (found in ecoli)

What part of CRISPR is used by humans?

CAS9

What is the CAS9?

CAS9 is a CRISPR associated protein/enzyme that contains both helicases and nucleases. These unwind and cut the DNA at certain spots. This does the gene editing.

Nucleases

Enzymes that naturally cut DNA

Helicases

Enzymes that unwind and expose DNA sequence

gRNA

Manipulated RNA that targets DNA of interest and provides new healthy DNA to be replaced after cut by CAS9

PAM

A guide that tells the nucleases where to cut the DNA and interrupt the sequence

Target Gene

The unhealthy gene

CAS9 Diagram

IVF (in vitro fertilization)

A procedure in which gametes are fertilized in a dish in the laboratory, and the resulting zygote is implanted in the uterus for development.

Zygote

A diploid cell resulting from the fusion (fertilization) of two haploid gametes (first cell of embryo)

Embryo

The early stages of development during the growth of a multicellular organism (2-8 weeks after fertilization)

Uses of PGD (preimplantation genetic diagnosis)

- Look for chromosomal abnormalities

- Sex of fetus

How does PGD work?

1-2 cells are extracted from the embryo and analyzed through the use of a karyotype

Reproductive Endocrinologist

A surgical subspecialty of obstetrics and gynecology that trains physicians in reproductive medicine addressing hormonal functioning as it pertains to reproduction as well as the issue of infertility.

Andrologist

The medical specialty that deals with male health, particularly relating to the problems of the male reproductive system and urological problems that are unique to men.

Embryologist

A fertility specialist that helps to create viable embryos to either be used in IVF right away or to be frozen for later use.

Why was Dolly so important?

She was the first successfully cloned mammal using the nucleus of somatic cells.

Therapeutic Cloning

The cloning of human cells by nuclear transplantation for therapeutic purposes, such as the generation of embryonic stem cells to treat disease.

Reproductive Cloning

Using a somatic cell from a multicellular organism to make one or more genetically identical individuals.

How does cloning work?

A mature somatic cell is taken from a patient and the nucleus is inserted into a donor egg. After the egg has developed, the embryo will either be implanted into a mother to fully develop or will grow to 100 cells (blastocyst) and then be destroyed for specific stem cell use.

Process of Gene Cloning (Basic)

A bacterial plasmid is altered through the use of restriction enzymes, recombinant DNA, ligase, and other tools. These are then grown with other bacteria and through conjugation are spread through a colony. These reproduce the integrated gene through protein synthesis and can then be used for human purposes.

Specific Use for Each Cloning

Gene: Used for the synthesis and mass production of a specific gene (vitamins, hormones, antibiotics)

Therapeutic: Used to make cells that can be inserted to produce the needed proteins

Reproductive: Used to make an exact copy of an entire organism