AP biology chapter 20- Biotechnology

DNA sequencing

Relatively short DNA fragments can be sequenced by the dideoxy chain termination method,

• Modified nucleotides called dideoxyribonucleotides (ddNTP) attach to synthesized DNA strands of different lengths

Gene cloning

Most methods for cloning pieces of DNA in the laboratory share general features, such as the use of bacteria and their plasmids; taking a gene from one source and putting it into another; involves using bacteria to make multiple copies of a gene

Plasmids

• are small circular DNA molecules that replicate separately from the bacterial chromosome

• Cloned genes are useful for making copies of a particular gene and producing a protein product

• foreign DNA is inserted into here and the recombinant is inserted into a cell, the reproduction in the bacterial cell results in cloning

Restriction enzymes

• cut DNA molecules at specific DNA sequences called restriction sites.

• it usually makes many cuts, yielding restriction fragments

The most useful restriction enzymes cut DNA in a staggered way, producing fragments with “sticky ends.”

gel electrophoresis

an indirect method of rapidly analyzing and comparing genomes; this technique uses a gel as a molecular sieve to separate nucleic acids or proteins by size, electrical charge, and other properties

• A current is applied that causes charged molecules to move through the gel

Molecules are sorted into “bands” by their size

Sticky ends

can bond with complementary sticky ends of other fragments

DNA ligase

is an enzyme that seals the bonds between restriction fragments

Restriction Fragment Analysis

• DNA fragments produced by restriction enzyme digestion of a DNA molecule are sorted by gel electrophoresis
  

• can be used to compare two different DNA molecules, such as two alleles for a gene if the nucleotide difference alters a restriction site

Polymerase Chain reaction

can produce many copies of a specific target segment of DNA

• A three-step cycle—- heating, cooling, and replication; brings about a chain reaction that produces an exponentially growing population of ideentical DNA molecules

• The key to PCR is an unusual, heat-stable DNA polymerase called Taq polymerase

Cloning Vector

• In gene cloning, the original plasmid is called a ____________

• 
  it is a DNA molecule that can carry foreign DNA into a host cell and replicate there

expression vector

• To overcome differences in promoters and other DNA control sequences, scientists usually employ this a cloning vector that contains a highly active bacterial promoter

electroporation

• One method of introducing recombinant DNA into eukaryotic cells is ______ applying a brief electrical pulse to create temporary holes in plasma membranes

• Alternatively, scientists can inject DNA into cells using microscopically thin needles

• Once inside the cell, the DNA is incorporated into the cell’s DNA by natural genetic recombination

Nucleic Acid Hybridization

If we have a gene we cloned and we want to know where it is expressed we can use __________

• Create a probe of complementary sequence that is single stranded

• Use a fluorescent tag on the probe to find where its hybridizing at

• If done in an embryo it allows us to study the mRNA in place (in situ)

Complementary DNA (cDNA)

• is made by cloning DNA made in vitro(glass) by reverse transcription(mRNA-DNA) of all the mRNA produced by a particular cell

A cDNA library represents only part of the genome—only the subset of genes transcribed into mRNA in the original cells

DNA microarray assays

scientists use this to measure expression of thousands of genes at one time; compare patterns of gene expression in different tissues, at different times, or under different conditions

Cas-9(CRISPR)

A special gene editing tool

SNPs(single nucleotide polymorphisms)

Genetic markers; occur on average ever 100-300 Base pairs; can be detected by PCR

• Any SNP shared by people affected with a disorder but not among unaffected people may pinpoint the location of the disease-causing gene

Organismal cloning

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

Totipotent cell

• is one that can generate a complete new organism (must dedifferentiate)

• Plant cloning is used extensively in agriculture

nuclear transplantation

• the nucleus of an unfertilized egg cell or zygote is replaced with the nucleus of a differentiated cell

stem cell

a relatively unspecialized

d cell that can reproduce itself indefinitely and differentiate into specialized cells of one or more types(plorypotent)

differentiate

the process where unspecialized cells become specialized, acquiring specific structures and functions through changes in gene expression and cellular activity

iPS cells (induced pluripotent cells)

these cells can be used to treat some diseases and to replace nonfuctional tissues

Gene Therapy

iI the alteration of an afflicted individuals genes; holds great potential for treating disorders traceable to a single defective gene

Vectors

are used for delivery of genes into types of cells

GLEEVAC

• s a small molecule that inhibits overexpression of a specific leukemia-causing receptor (CML patients only)

• Pharmaceutical products that are proteins can be synthesized on a large scale

Transgenic

these types of animals are made by introducing genes from one species into the genome of another

Short tandem repeats (STRS)

a type of genetic marker, which are variations in the numbers of repeats of Specific DNA sequences

Genetic profile

An individual’s unique DNA sequence can be obtained by analysis of tissue or body fluids