Ap Bio Outline

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- Cloning Plants: Single-Cell Cultures

- Organismal cloning: cloning produces one or more organisms genetically identical to “parent” that donated the single cell

- Genomic equivalence: concept that all cells of an organism have same genes

- Totipotent: cell with potential of giving rise to all specialized cell types of the organism

- Cloning Animals: Nuclear Transplantation

- Differentiated cells from animals generally do not divide in culture

- Nuclear transplantation: remove nucleus of an unfertilized or fertilized egg and place it with the nucleus of a differentiated cell

- Potential of a transplanted nucleus to direct normal develpment was inversely related to the age of the donor (older donor → lower percentage of normal development)

- Reproductive Cloning of Mammals

- Dolly reproductive cloning

- Mammary cell donor donates cells, one of which fuses with to an enucleated egg cell from an egg cell donor

- Egg cell with nucleus from mammary cell grows in culture

- Early embryo is implanted in uterus of a third sheep (surrogate mother)

- Dolly is genetically identical to mammary cell donor (differs from that of egg donor and surrogate mother)

- Out of several hundred implanted embryos, only one was successful

- Dolly later died prematurely, reflecting possible incomplete reprogramming of the original transplanted nucleus

- Cloned animals of same species do not always look or behave identically

- First cloned cat (calico cat) → Carbon Copy (CC)

- Color and pattern is different from single female parent because of random X chromosome inactivation

- Identical human twins are naturally occuring “clones” → always slightly different

- Problems Associated with Animal Cloning

- Low efficiency of cloning and high incidence of abnormalities

- In nuclei of fully differentiated cells, a small subset of genes is turned on an expression of rest is repressed (regulation is result of epigenetic changes in chromatin)

- DNA in cells from cloned embryos often have more methyl groups than does DNA in equivalent cells from normal embryos of same species

- Reprogramming of donor nuclei requires chromatin restructuring, which occurs incompletely during cloning procedures

- Misplaced methyl groups in DNA of donor nuclei may interfere with pattern of gene expression necessary for normal embryonic development

- Stem Cells of Animals

- Stem cell: relatively unspecialized cell that can both reproduce itself indefinitely and differentiate into specialized cells of one or more types

- Can be isolated from early embryos at blastula/blastocyst stage (ES cells)

- Adult stem cells cannot give rise to all cell types in the organism, though they can generate multiple types

- ES cells are pluripotent (capable of differentiating into many different cell types)

- Therapeuthic cloning: cloning to produce ES cells to treat diseases

- Induced pluripotent (iPS) cells: differentiated cells transformed into ES cells by using retoviruses to introduce extra cloned copies of four “stem cell” master regulatory genes

- Cells from patients suffering from diseases can be programmed to become iPS cells, which can be used to study the disease

- Patient’s own cells can also be reprogrammed into iPS cells and then used to replace nonfunctional tissues

20.4 The practical applications of DNA technology affect our lives in many ways

- Medical Applications

- Diagnosis and Treatment of Diseases

- RT-PCR is often the best way to detect an otherwise elusive infective agent

- Medical scientists can now diagnose hundreds of human genetic disorders by using PCR with primers that target the genes associated with these disorders

- Amplified DNA product is sequenced to reveal the presence or absence of the disease-causing mutation

- Inidividuals afflicted with such diseases can often be identified before the onset of symptoms, even before birth

- Individuals can be tested for SNP that indicates abnormal allele

- Human Gene Therapy

- Gene therapy: introducing genes into an afflicted individual for terapeutic purposes

- In theory, normal allele of defective gene could be inserted into somatic cells of tissue affected by the disorder

- For gene therapy of somatic cells to be permanent, cells the receive normal allele must be ones that multiply throughout the patient’s life

- Gene threapy using a retoviral vector (in bone marrow cells)

- Insert RNA version of normal allele into retrovirus

- Let retrovirus infect bone marrow cells that have been removed from the patient and cultured

- Viral DNA carrying the normal allele inserts into chromosome

- Three of the patients of the SCID gene therapy trials developed leukemia

- Two factors that may have contributed to development of leukemia: insertion of the retroviral vector near a gene involved in the proliferation of blood cells and an unknown function of the replacement gene itself

- Gene therapy provokes technical