Gene Technology Flashcards

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1
Q

Genome

A

Complete set of genes in a cell

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2
Q

Recombinant DNA

A

A cell having two or more sources of DNA
Possible as genetic code is universal,degenerate and non-overlapping

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3
Q

5 steps in recombinant DNA technology

A

Isolation of genes, insertion, transformation, identification and growth

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4
Q

1- isolating a copy of specific DNA fragment
What are the different methods

A

Gene machine
Reverse transcription
restriction endonucleases

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5
Q

1-using reverse transcriptase

A

Isolated mRNA from donor cell
Add dna nucleotides and reverse transcriptase
Free dna nucleotides bind to single stranded mRNA via complimentary base pairing
Reverse transcriptase joins dna nucleotides together to form single standard cDNA molecule
Add dna polymerase to make cDNA double stranded

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6
Q

Advantages of using reverse transcriptase

A

mRNA is easy to obtain
Using mRNA means introns removed whereas genes contain both introns and exons

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7
Q

2-using restriction endonuclease

A

They hydrolyse dna at specific base sequence (either side of gene)
Regcognition sequence is usually palindromic -based pair is same whether read forwards or backwards
Cut made is blunt(used in pcr and gel electrophoresis)
Or sticky(used in transformation)

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8
Q

C-gene machine

A

Enter the desired sequence of nucleotides into a computer
Creates small sections of overlapping single strands of nucleotides
Joined to make dna
Pcr used to amplify and make double stranded

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9
Q

Advantages of the gene machine

A

Artificial genes are easily transcribed and translated by prokaryotes as they have no introns in their dna

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10
Q

What is a vector

A

DNA carrier

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11
Q

Insertion of genes into a vector

A

cut vector using the same restriction enzyme that was used to isolated dna fragments
-Produces complimentary sticky ends between ends of dna fragment and cut ends of the vector dna
-Target dna fragment anneals to vector dna by complimentary base pairs between their sticky ends
-DNA ligase used to join them at sugar phosphate backbone and forms phosphodiester bonds

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12
Q

Transformation

A

Heat shock/ electric current used to make temporary holes in cell curvaceous membrane
Bacterium takes up vector plasmid
Recombinant organism

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13
Q

Identifying transformed host cells

A

not all vectors take up target dna to become recombinant
Marker genes are added
Fluorescence - fluoresces when exposed to uv light
Enzyme markers - no colour change indicted required gene has been inserted

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14
Q

Identifying transformed bacteria using antibiotic resistance genes

A

Cells that didn’t take up plasmid are killed by both types of antibiotics
Cells that took up original plasmid are resistant to both types of antibiotics
Cells that took up the transformed plasmid are resistant to one antibiotic

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15
Q

Cloning in vitro via pcr

A

Pcr used to amplify
Several stages that automatically repeat
1 seperation - strands separate at 95 c
2 annealing -addition of primers ,primers bond 50c
3 synthesis - makes 2 dna fragments 1old 1 new

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16
Q

Primer

A

Short pieces of single stranded dna with complimentary base sequences to dna fragments

17
Q

Benefits of gene tech

A

Develop medical applications
Better understanding of biological processes

18
Q

Concerns with gene tech

A

Introducing herbicide resistant genes to crops
could result in transfer to wild plants
Which could become resistant to herbicides

19
Q

Somatic and germ line gene therapy

A

Somatic - dna transfer to our normal body tissue
Germ line-dna transfer to cells that produce eggs or sperm

20
Q

Limitations of somatic and germ line gene therapy

A

Not all cells take up new dna
Multiple treatments may be needed

21
Q

Objections of gene therapy

A

Tech is imperfect
Denial of human rights
Potential abuse

22
Q

Genetic screening using dna probes and dna hybridisation

A

-determine nucleotide sequence of the mutant allele that you’re trying to find
Create comp dna probe and add a marker
-many dna probes made using pcr
-dna with suspected alle is heated to separate both strands
-seperated strands are cooled in a mixture of dna probes
If strand has mutant allele probe will bind to it as bases are complimentary
-dna is washed to remove unattached probe
-remaining hybridised dna will be fluorescently binded
-dye is detected by shining light causing fragment to fluoresce

23
Q

Gene probe

A

Short, single stranded dna molecule with complimentary base sequence to dna fragment

24
Q

Genetic fingerprinting and VNTR’s

A

Uses the region of dna between genes called VNTR’s
Each person has 2 copies of each VNTR on each homologous chromosome
1 from each parent
Difference in length in VNTR can be identified by gel electrophoresis

25
Q

DNA fingerprinting

A

-extract dna from the nucleus of cells
-pcr amplifies VNTR regions
restriction enzymes are used to cut dna into fragments
-must make blunt ends
-separate dna fragments using gel electrophoresis
-put dna in well and apply electrical current
-make fragments single stranded by soaking in alkali solution
-add radioactive gene probes with comp base sequences to VNTR
-identify base sequence using x ray film

26
Q

Using liposomes as vectors

A

Using liposomes as vectors
Lipid droplets which can cross phospholipid bilayer and releases target dna into the cell
However dna does not move into the nucleus so new daughter cells will not have the functional gene