EX3 DNA TECHNOLOGY

• -DNA Tech: manipulation of
• genetic material, even moving genes from one organism to another

• -Recombinant DNA: a DNA
• molecules that derives from two or more sources

• -Genetically modified organism:
• organism with one or more genes introduced using DNA technology

• -Transgenic organism: A GMO
• which has genes from more than one organism

Reaction enzyme: proteins that cleave DNA at restriction sites Sticky Ends: any DNA cut by a particular restriction enzyme can stick to any other cut by the same enzyme

A genetic structure in a cell that can replicate independently of the chromosome, typically a small circular DNA strand in the CYTOPLASM of a bacterium

• 1.    
• getting
• smaller pieces of DNA to wok with restriction enzymes

• 2.    
• separating
• fragments: gel electrophoresis

• 3.    
• find
• a particular sequence in the genome: nucleic acid probes

• 4.    
• making
• many copies of the DNA: cloning by biological vector or polymerase chain
• reaction

Separating molecules by size, charge etc -measures movement based on a standard -placed in gel and move by electrical charge

• 1.    
• vector
• such as bacteria, yeast, cell culture

• -gene cleaved out of original genome using a particular
• restriction enzyme

• -gene inserted into bacteria plasmid (vector) cut with the
• same enzyme to transfer (recombinant DNA) by shaking them together

•        2.  
• Polymerase chain reactions (PCR)

•                   -clones of DNA sequence
• without a biological vector

• 1.    
• special
• engineering plasmid and DNA of interest cut with same restriction enzyme and
• mixed together, some combine

• 2.    
• mixtures
• is exposed to a biological vector such as yeast or bacteria.  Some vectors pick up DNA

• 3.    
• individual
• vectors are screened to identify those which picked up a plasmid which
• contained the DNA of interest

• 4.    
• These
• selected vectors are allowed to reproduce making more copies of the DNA of
• interest

PCR produces copies from one sample

•                   -primer(short
• sequence of dna to be amplified)

•                   -target
• sequence (dna)

•                   -dna
• pol (heat-stable version-TAQ Polymerase)

•                   -nucleotides
• monomers

 

STEPS:

•                   -heated
• to 95 C, so dna denatures to single strands

•                   -cools,
• primers attach to their complimentary sequence

•                   -dna
• pol adds nucleotides to finish complimentary strand

•                   -more
• rounds yield more copies

Nucleic acid probe: finding the correct gene

•                   -single
• stranded sequence of interest is labeled (dye, radiation)

•                   -when
• mixed with the sample, it sticks to its compliment

•                   -any
• single stranded sequence

• ex. 
• CCGGTTAA PROBE à DNA sequence labeled

                                                             |CCGGTTAA|

                                                      TAT|CCGGTTAA|CTGAGA

• -based on
• highly variable non-coding sequences

• -cut with
• restriction enzymes

• -many
• radioactive probes used to determine which fragments are present

• -compared to
• suspects

• -very low
• chance of error

• -based on
• single tandem repeats (STR’s) non-coding DNA more variable than coding DNA

• The study of
• entire genomes

• About 4000
• species sequenced (mostly prokaryotes)

Human genome:

• Protein coding
• DNA 1-15%

• Non coding DNA
• 98.5-99%

• Non-coding
• DNA:

• -24% introns
• and regulatory sequence

• -15% structural
• DNA around centromeres or ends (telomoeres); mostly simple sequence repeats

• -44%
• transposable elements (jumping genes)

• Humans are
• more complex with approx. 100,000 genes

• Passing of a
• chemical fragment called methyl group from one molecule to another.

• It silences a
• gene