Cloning Lecture

Cloning Objectives • Define cloning. • Identify the history of cloning. • Analyze the different types of cloning. What is Cloning? • Animal cloning is an assisted reproductive technology similar to artificial insemination, embryo transfer, and in vitro fertilization. • The current technique used for animal cloning is somatic cell nuclear transfer (SCNT). • There are no published US guidelines for unique requirements regarding the care and use of animal clones in research. • The care and use of animal clones in research does not differ from care provided for conventional animals to assure good animal welfare and animal well-being. In addition, because the progeny of animal clones are not clones, clearly progeny do not require special consideration. Cloning • Reproducing a genetically identical offspring Introduction to Cloning • The possibility of human cloning, raised when Scottish scientists at Roslin Institute created the muchcelebrated sheep "Dolly" (Nature 385, 810-13, 1997), aroused worldwide interest and concern because of its scientific and ethical implications. • The feat, cited by Science magazine as the breakthrough of 1997, also generated uncertainty over the meaning of "cloning" --an umbrella term traditionally used by scientists to describe different processes for duplicating biological material. Types of Cloning • The following three types of cloning technologies will be discussed: – (1) recombinant DNA technology or DNA cloning – (2) reproductive cloning – (3) therapeutic cloning. Recombinant DNA Technology or DNA Cloning • • • • The terms "recombinant DNA technology," "DNA cloning," "molecular cloning," and "gene cloning" all refer to the same process: the transfer of a DNA fragment of interest from one organism to a self-replicating genetic element such as a bacterial plasmid. The DNA of interest can then be propagated in a foreign host cell. This technology has been around since the 1970s, and it has become a common practice in molecular biology labs today. Scientists studying a particular gene often use bacterial plasmids to generate multiple copies of the same gene. Plasmids are self-replicating extra-chromosomal circular DNA molecules, distinct from the normal bacterial genome (see image to the right). Plasmids and other types of cloning vectors were used by Human Genome Project researchers to copy genes and other pieces of chromosomes to generate enough identical material for further study. Recombinant DNA • To "clone a gene," a DNA fragment containing the gene of interest is isolated from chromosomal DNA using restriction enzymes and then united with a plasmid that has been cut with the same restriction enzymes. – When the fragment of chromosomal DNA is joined with its cloning vector in the lab, it is called a "recombinant DNA molecule." – Following introduction into suitable host cells, the recombinant DNA can then be reproduced along with the host cell DNA. Recombinant DNA Cont. • Plasmids can carry up to 20,000 bp of foreign DNA. • Besides bacterial plasmids, some other cloning vectors include viruses, bacteria artificial chromosomes (BACs), and yeast artificial chromosomes (YACs). • Cosmids are artificially constructed cloning vectors that carry up to 45 kb of foreign DNA and can be packaged in lambda phage particles for infection into E. coli cells. BACs utilize the naturally occurring F-factor plasmid found in E. coli to carry 100- to 300-kb DNA inserts. • Bacteria are most often used as the host cells for recombinant DNA molecules, but yeast and mammalian cells also are used. Reproductive Cloning • Dolly, the first mammal to be cloned from adult DNA, was put down by lethal injection Feb. 14, 2003. • Reproductive cloning is a technology used to generate an animal that has the same nuclear DNA as another currently or previously existing animal. – Dolly was created by reproductive cloning technology. – By a process called "somatic cell nuclear transfer" (SCNT), scientists transfer genetic material from the nucleus of a donor adult cell to an egg whose nucleus, and thus its genetic material, has been removed. – The reconstructed egg containing the DNA from a donor cell must be treated with chemicals or electric current in order to stimulate cell division. – Once the cloned embryo reaches a suitable stage, it is transferred to the uterus of a female host where it continues to develop until birth. Reproductive Cloning • Dolly or any other animal created using nuclear transfer technology is not truly an identical clone of the donor animal. – Only the clone's chromosomal or nuclear DNA is the same as the donor. – Some of the clone's genetic materials come from the mitochondria in the cytoplasm of the enucleated egg. Mitochondria, which are organelles that serve as power sources to the cell, contain their own short segments of DNA. – Acquired mutations in mitochondrial DNA are believed to play an important role in the aging process. Therapeutic Cloning • Therapeutic cloning, also called "embryo cloning," is the production of human embryos for use in research. • The goal of this process is not to create cloned human beings, but rather to harvest stem cells that can be used to study human development and to treat disease. • Stem cells are important to biomedical researchers because they can be used to generate virtually any type of specialized cell in the human body. Stem cells are extracted from the egg after it has divided for 5 days. • The egg at this stage of development is called a blastocyst. • The extraction process destroys the embryo, which raises a variety of ethical concerns. • Many researchers hope that one day stem cells can be used to serve as replacement cells to treat heart disease, Alzheimer's, cancer, and other diseases. Advantages of Cloning • • • • Utilize good genetics Make steers into bulls Clone for insurance Get another good animal Disadvantages of Cloning • • • • Expensive Will animals be normal? Takes many times to clone Dangerous to humans?