5-2a: Cloning

• How are mammals cloned?• What is the difference between reproductive

cloning and therapeutic cloning?

Hello, Dolly!

Dolly and her surrogate motherRoslin Institute

Dolly and her lamb, BonnieRoslin Institute

Mammalian cloning via somatic cell nuclear transfer

Anne McLaren, 2001 Nature 414, 129-131 doi:10.1038/35102194

Why has cloning mammals become possible now?• In vitro fertilization technology has come of

age. Louise Brown the first “test tube baby” was born in the 1970s.

• Nuclear transfer into human eggs is possible for males with defective sperm.

• Embryo transfer is routinely used for reproduction of livestock (cattle)

How easy (efficient) is animal reproductive cloning?

Dolly was the only lamb from 277 fused eggs Many animal species have been cloned.

Tetra – rhesus monkey

CopycatIdaho Gem mule

http://www.businesspundit.com/20-animals-that-have-been-cloned/

Are cloned animals healthy?Dolly gave birth to a healthy lamb named Bonnie.Dolly suffered prematurely from arthritis and other ailments of old age.Dolly’s telomeres (ends of chromosomes) were shorter than for other sheep of comparable age.

Complications in reproductive cloning

• Cloned offspring syndrome– Placental abnormalities– Fetus too large– High rate of spontaneous abortions– Early death

• Shorter telomeres?• Genes not imprinted properly

– Genes that control placental growth, fetal growth inactivated in maternal chromosomes

Therapeutic cloning produces embryonic stem cells

Embryonic stem cell colony

5-2b: Stem Cells

• What is the difference between embryonic stem cells and adult stem cells?

• What legal and ethical issues revolve around cloning and stem cell technologies?

Stem cells: undifferentiated cells that can keep dividing to replenish various differentiated cell populations

• Totipotent – embryonic stem cells• Pluripotent – bone marrow stem cells• Oligopotent – hematopoietic stem cells• Multipotent – erythropoietic stem cells• Unipotent – reticulocytes

Some adult stem cells

mammalan development in vitro from egg to blastocyst

http://yaolab.stanford.edu/research/preimplantation.html

Human development through blastocyst implantation

© 2001 Terese Winslow, http://stemcells.nih.gov/info/scireport/appendixA.asp

Generating embryonic stem cells for regenerative medicine

Klimanskaya I, N Rosenthal, R Lanza 2008, Nature Reviews Drug Discovery 7, 131-142 doi:10.1038/nrd2403

Alternatives to embryonic stem cells?• Induced pluripotent stem cells

– Aoi et al. 2008, Generation of Pluripotent Stem Cells from Adult Mouse Liver and Stomach Cells, Science 321:699-702

– Retroviral vectors used to transduce 4 transcription factors, incl. c-Myc (proto-oncogene)

– ES-like cells in gene expression profile, pluripotency (form tumors containing different cell types, form all 3 major tissue types, germline transmission)