Asexu. Figure 6 A small glass tube, called a micropipette, is used to remove the nucleus from a cell and later introduce a new nucleus.

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1 While plant cloning experiments were being conducted, Robert Briggs and Thomas King were busy investigating nuclear transplants in frogs. Working with the common grass frog, the scientists extracted the nucleus from an unfertilized egg cell by inserting a fine glass tube, or micropipette, into the cytoplasm and sucking out the nucleus (Figure 6). A cell without a nucleus is referred to as enucleated. erm d Asexu a l Repro ducti o n: 2. Clo ning Figure 6 A small glass tube, called a micropipette, is used to remove the nucleus from a cell and later introduce a new nucleus. Next, the nucleus of a cell from a frog embryo in the blastula stage of development was removed and inserted into the enucleated cell. The egg cell with the transplanted nucleus began to divide much like any normal fertilized egg cell. In later trials, the cell with the transplanted nucleus occasionally grew into an adult frog. Not surprisingly, the adult frogs displayed the characteristics from the transplanted nucleus. Careful analysis proved that the adults were clones of the frog that donated the nucleus (Figure 7). However, different results were obtained when the nucleus was taken from cells at later stages of development. A nucleus that can bring a cell from egg to adult is referred to as totipotent, but not all nuclei are totipotent. For example, the nucleus from cells in a later stage, called the gastrula stage, did not bring the enucleated egg from the single-cell stage to the adult. If cell division occurred at all, it did not progress as far as for eggs that received a blastula nucleus. The difference is that the nucleus of a cell in the gastrula stage of development, unlike a cell in the earlier blastula stage, has specialized. As cells begin to specialize, a regulatory mechanism must turn off some of the genes that allow cell division. Figure 8 Nature s clones. The left photograph shows identical twins. They originate from a single fertilized egg that

2 What is Cloning? Is the process that produces identical copies of genes, cells or organisms

3 Types of Cloning 1. Gene Cloning is the practice of manipulating DNA to make multiple copies of a gene or a piece of DNA in foreign cells. Practical Applications ~ mass production of insulin 2. Therapeutic Cloning is the production of genetically identical cells. Practical Applications ~ organ/tissue growing 3. Reproductive Cloning is the process of producing genetically identical organisms. ex: Dolly the Sheep. Practical Application ~ repopulate endangered species

4 Gene Cloning Bacteria have a large circular chromosome as well as many smaller circular structures called plasmids. These plasmids are an important tool in gene splicing. 1 µm

5 Gene Cloning Bacteria have a large circular chromosome as well as many smaller circular structures called plasmids. These plasmids are an important tool in recombinant DNA techniques. 1 µm

6 Gene Cloning 1. Human DNA is cut into fragments using a enzyme.

7 Gene Cloning Human cell insulin gene DNA 1. Human DNA is cut into fragments using a enzyme. Many fragments will be made, but one will have the insulin gene.

8 Gene Cloning 1. Human DNA is cut into fragments using a enzyme. Many fragments will be made, but one will have the insulin gene. 2. Plasmids are cut with the same enzyme as in step1.

9 Gene Cloning complimentary ends The antibiotic resistance gene will be important in a later process. antibiotic resistance gene 1. Human DNA is cut into fragments using a enzyme. Many fragments will be made, but one will have the insulin gene. 2. Plasmids are cut with the same enzyme as in step1. The plasmids also contains an antibiotic resistance gene.

10 Gene Cloning 3. Mix the DNA fragments, the cut plasmids, to produce recombinant DNA plasmids.

11 Some of these recombinant plasmids will contain the insulin gene, and some will not. 3. Mix the DNA fragments, the cut plasmids, to produce recombinant DNA plasmids.

12 Gene Cloning 4. Through transformation, recombinant plasmids enter bacterial cells. As the bacteria divide billions of copies of the recombinant plasmids are made. Some of these bacteria will made insulin

13 Gene Cloning 5. A process called hybridization is used to identify the bacterial colonies with the desired DNA.

14 Transgenic Organisms From our knowledge of Cloning, Geneticists of developed techniques of inserting foreign DNA in plants and animals to produce Transgenic Organisms These are a type of Genetically Modified Organisms (GMO) For example: Spider silk, BT corn, etc.

15 Reproductive Strategies & Technologies

16 History... For thousands of years humans have used reproductive technologies for the development of their livestock and plants. Today we are able to use this knowledge to manipulate organisms genetically in new ways in Agriculture & Humans

17 Reproductive Technologies in Agriculture Farmers carefully select plants and animals with specific desirable traits for breeding ~ Selective Breeding Technologies that help with this: 1. Artificial Insemination: the process by which sperm are collected and concentrated before being introduced into the female s reproductive system 2. Embryo Transfer: The process by which an egg that has been fertilized artificially is transferred into a recipient female s uterus.

18 Reproductive Technologies for Humans There are a number of reproductive technologies that are now available for couple who are not able to conceive a child These are often referred to as Assisted Reproductive Technologies (ART) It is important to note that as these ART techniques led to cures, treatments and new inventions that they also stir controversy ~ Catholicism

19 Assisted Reproductive Technologies (ART) ~ Sexual Reproduction 1. Artificial Insemination 2. In Vitro Fertilization (IVF) 3. Preimplantation Genetic Diagnosis (PGD)

20 1. Artificial Insemination The sperm of the desired male are collected and concentrated Concentrated sperm is then introduced into the woman s vagina This allows sperm from the woman s male partner or from an unknown source to be used in implantation i.e. sperm bank.

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22 2. In Vitro Fertilization Also known as IVF & babies conceived this way are known as Test tube babies This is beneficial for woman who have blocked Fallopian tubes. Immature eggs are retrieved from that woman Eggs are combined with sperm in Laboratory glassware (In Vitro) where sperm fertilizes egg The developing embryo is then placed in the uterus

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26 3. Preimplantation Genetic Diagnosis Families who have a history of genetic disorders in their family may use this This is used soon after IVF fertilization so that Doctors may diagnose genetic disorders (before implanted in uterus) embryos are analyzed for the presence of a genetic disorder and only the healthy ones are implanted This technique is also used to engineer genetic matches in another sibling i.e. able to donate umbilical cord blood that has stem cells

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