Biotech Test 2

Front 1

How are nucleotides held together?

Back 1

hydrogen bonds.

Front 2

Antiparallel

Back 2

The nucleotides in each strand are oriented in opposite direction of the other strand

Front 3

Semiconservative replication

Back 3

making two daughter strands from one single strand.

Front 4

Variations in DNA molecules

Back 4

The # of DNA strands in the cells of an organism (chromosomes)

The length in base pairs

the # and type of genes (nucleotide sequences that code for protein production) and noncoding region the shp of the DNA strands (circular or linear)

Front 5

Bacteria DNA

Back 5

contains one long, circular DNA molecule that is supercoiled. The DNA molecule is rather small and only contains several thousand of genes.

Some bacteria have small rings of DNA floating in their cytoplasm → plasmid: only contains a few genes that code for proteins which can offer some additional characteristic needed under extreme conditions

Front 6

R Plasmid

Back 6

Plasmids that have been modified, often containing antibiotic resistance genes.

Front 7

Transformed Cells

Back 7

cells that have taken up foreign DNA and expressed the newly acquired gene

Front 8

Operon

Back 8

a section of prokaryotic DNA consisting of one or more genes and their controlling elements.

Front 9

Enzymes involved in DNA replication

Back 9

Helicase- Separating the double stranded DNA. Topoisomerase- relieves tension from helicase. Unclips one strand, unwinds it, and puts it back.(RNA) Primase- Adds a small segment of RNA prior to the DNA from polymerase. Demonstrates that RNA developed first.Polymerase- Copying DNA.Ligase- Ties fragments of DNA together.

Front 10

DNA coiling

Back 10

DNA is coiled around Histone proteins.

Front 11

Make up of Viruses

Back 11

A protein coat encasing DNA or RNA.

Front 12

Gene Therapy

Back 12

the process of treating a disease or disorder by replacing the dysfunctional gene with a functional one.

Front 13

Recombinant DNA

Back 13

DNA comprised from two or more organisms

Recombinant DNA technologies are the methods used to create new DNA molecules by piecing together different DNA molecules

Front 14

Genetic Engineering

Back 14

Manipulating genetic information to produce new characteristics or to transfer characteristics between organisms.

Front 15

Probe

Back 15

A small complimentary strand of DNA that hybridizes with a larger strand and, because it is fluorescent or radioactive, it can locate specific genes.

Front 16

Southern Blotting

Back 16

A method of locating specific DNA using probes

Isolate the DNA

Run a gel - chromatography - separating DNA, finding individual segments of DNA

DNA fragments are separated by electrophoresis - chromatography

Fragments are transferred to a gel membrane

Probes are added to the surface

The membrane is observed to see which DNA fragments hybridized with the probes

Front 17

Components of PCR

Back 17

DNA polymerase, Template DNA, primers, dNTPs, Buffer.

Front 18

Primer construction

Back 18

a short DNA segment that creates binding site for the DNA polymerase

Optimally have a 50% G-C content

15 - 30 bps

Complementary to recognition sequence

Copies past gene to not cut it off

Do not self-anneal - primer dimers, hairpins

Front 19

Steps in PCR

Back 19

1. Denaturation - “melting” - separation of double helix at 95 degrees

2. Annealing - connecting - base pairing of a primer at 55 degrees

3. Extension - Polymerase acts to replicate DNA at 72 degrees

Front 20

Recombinant DNA Technology:

Back 20

Process of Artificially having an organism produce a protein.

Front 21

Restriction enzymes cutting

Back 21

Restriction Enzymes cut palindromic sequences

Front 22

Hybridize

Back 22

Complementary base-pairing

Front 23

General Spectrophotometry

Back 23

A process using visible or UV light to measure the transmission and absorption of light in a sample.

Front 24

Transmittance

Back 24

The passing of light through a sample.

Front 25

Absorbance

Back 25

The amount of light absorbed by the sample (the amount of light that does not pass through or reflect off the sample). Absorbance is when a substance takes in and uses the light as an energy source, or when electrons are excited. The sample is USING ENERGY.

Front 26

Concentration vs. Absorbance vs. Transmission

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As the concentration of a sample increases, so does absorbance; when the concentration of a sample decreases, absorbance decreases as well. As transmittance goes up, concentration of sample is lower; as transmittance goes down, concentration of sample is higher.

Front 27

Uses of General Spectrophotometry

Back 27

determining the identity of a substance, and in determining if a substance is present in a solution/sample.

Front 28

Components of a Spec

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· Deuterium lamp (UV for colorless proteins and Nucleic Acids) or Tungsten lamp (White or visible light).

· Prisms that allow one wavelength to go through.

Sample holders and displays.

Front 29

Visible Light

Back 29

has several colors, expressed through ROYGBIV.

· When a substance is hit with visible light, it appears to an observer to be the color of the light it reflects.

Front 30

Wavelength vs. Frequency vs. Energy

Back 30

The shorter the wavelength, the higher the frequency. High frequency waves have more energy that low frequency ones. Ex: radio waves have less energy than gamma waves.

Front 31

Blank

Back 31

a control; something that should have no absorbance, and is used to set the 0 au point.

· If, for example, you want to measure bacteria in media, use the media without the bacteria as a blank; generally, use whatever medium (media, water) you put your samples in.

Front 32

Optical Density

Back 32

· Absorbance is also known as Optical Density (O.D.).

· In bacteria, O.D. 600 = 1.0 = 8 x 10^8 cells/mL; induce at 0.7 au (O.D. 0.7). Proteins absorb at 280nm.

Front 33

Purity and Concentration

Back 33

· Purity- if reading is below 1.5, it is bad; if it is 1.8-2., it is very good.

DNA Concentration Formula:

50 ug/mL X ug/mL

------------------- = -----------

1 au at 260 nm The Absorbance of sample at 260 nm

DNA Purity Formula:

absorbance (au) at 260 nm

----------------------------------- = The purity of the sample

absorbance (au) at 280 nm

Front 34

DNA Uptake/ Competence

Back 34

Cells in a state of need will uptake DNA, can update modified DNA that when expressed will produce a DNA product.

Competent cells can uptake DNA

Front 35

Artificial cometence

Back 35

CaCl, cold

heat shock for 45 seconds in 42oC - stresses cells so they are in a state of need and will accept DNA

Electroporation:

shock cells so there are holes in the membrane that DNA can enter through

Front 36

Conjugation

Back 36

Bacteria sex. Horizontal gene transfer. *Requires direct contact* Donor cell: F plasmid (makes pilus, fertility factor). Nicked (cut) plasmid. One strand goes and one stays.

Front 37

Transduction

Back 37

transfer of DNA by bacteriophages - viruses that infect bacteria

no physical contact

viruses: nucleic acid and protein

Front 38

Transfection

Back 38

DNA into Eukaryotes

need to get DNA into nucleus of cell

doesnt happen in nature

Front 39

CFU calculations

Back 39

See sheet.