Genetics and Recombinant DNA BIT 120 Mitosis 46 chromosomes 23 pairs 22 pairs autosomes ( Chromosomes other than the X or Y sex chromosomes) 1 pair sex chromosomes: XX and XY
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Rr, round & wrinkled
Yy, yellow and green
Arrows indicate genes on the X chromosome for which there is no complement on the Y chromosome
1.DeletionHere, certain nucleotides are deleted, which affects the coding of proteins that use this DNA sequence. If for example, a gene coded for alanine, with a genetic sequence of C-G-G, and the cytosine nucleotide was deleted, then the alanine amino acid would not be able to be created
2. InsertionSimilar to the effects of deletion, where a nucleotide is inserted into a genetic sequence and therefore alters the chain thereafter. This alteration of a nucleotide sequence is known as frameshift
3.InversionWhere a particular nucleotide sequence is reversed, and is not as serious as the above mutations. This is because the nucleotides that have been reversed in order only affect a small portion of the sequence at large
4.SubstitutionA certain nucleotide is replaced with another, which will affect any amino acid to be synthesized from this sequence due to this change. If the gene is essential, i.e. for the coding of hemoglobin then the effects are serious, and organisms in this instance suffer from a condition called sickle cell anemia. - CAN BE SILENT MUTATION. CONSERVATION SUBSTITUTION, OR SUBSTITUTION
converts a GAG codon (for Glu) to a GTG codon for Val
abolishes a sequence (CTGAGG, which spans codons 5, 6, and 7) recognized and cut by one of the restriction enzymes.
DEAE dextran - an inert carbohydrate polymer (dextran) coupled to a positively charged chemical group (diethylaminoethyl -DEAE). DNA probably sticks to DEAE-dextran via its negatively charged phosphate groups.
Calcium phosphate - forms an insoluble precipitate with DNA. It was discovered that cells efficiently take up this precipitate. More efficient than DEAE dextran or many cell types and can be used for both transient and stable transfection. Not suitable for cells which grow in suspension culture.
Electroporation - Cells are concentrated, mixed with the DNA and placed in a small chamber with electrodes connected to a specialised power supply. A brief electric pulse is applied, which is thought to ‘punch holes’ in the cell membrane, enabling the cell to take up DNA.
Lipofection - (liposome-mediated gene transfer) several lipid- based methods have been developed in which DNA is encapsulated by synthetic lipid bilayers which resemble cell membranes. Liposomes are essentially spheres of synthetic membrane filled with DNA. These fuse spontaneously with cell membranes, releasing their contents into the cytoplasm.