biologia general 11 bioteecnologia - biologia.ucr.ac.crbiologia.ucr.ac.cr/profesores/garro...
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Biotecnología Prof. Bernal Gerardo Garro Mora
Escuela de Biología
UCR
Objetivos:
• Al finalizar la clase el estudiante será capaz de:
1. Entender la utilidad de los plásmidos, enzimas de restricción y ligasas en la formación del ADN recombinante
2. Explicar la técnica del PCR y describir su uso en la amplificación de porciones del genoma.
3. Describir los pasos cómo se obtiene la huella de ADN y su utilidad en ciencias forenses.
4. Entender cómo se desarrollan los cultivos genéticamente modificados.
5. Entender el estudio del genoma humano y la relevancia de sus resultados.
6. Conocer las aplicaciones de la genómica y sus implicaciones bioéticas.
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¿Qué entiende por ‘biotecnología’ e ‘ingeniería genética’?
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¿Qué es la biotecnología?
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Concepto de Biotechnología
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Concepto
de Biotechnología
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Colores de la biotecnología
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Algunas herramientas
de la ingeniería
genética
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Organismos transgénicos en la vida cotidiana
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plasmid
plasmid
bacterial
chromosome
(a) Bacterium
DNA
fragments
bacterial
chromosome
(b) Transformation with a DNA fragment
(c) Transformation with a plasmid
A DNA fragment is
incorporated into
the chromosome
bacterial
chromosome
The plasmid replicates
in the cytoplasm
1 micrometer
Transformation in Bacteria
Fig. 13-1
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The Life Cycle of a Typical Virus
Fig. 13-2
The virus releases itsDNA into the host cell; some viral DNA (red) may be incorporated intothe host cell’s DNA (blue)
The virus enters the host cell
Viral genes encode the synthesis of viral proteins and viral gene replication; some host cell DNA may attach to the replicated viral DNA (red/blue combination)New viruses assemble;
some host cell DNA is carried by recombinant viruses
The host cell bursts open, releasing newly assembled viruses; if recombinant viruses infect a second cell, they may transfer genes from the first cell to the second cell
viral proteinsrecombinant virus
virus
viral DNA
viral DNA
A virus attaches to a susceptible host cell
host
cell
host cell DNA
2
3
1
4
5
6
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PCR
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18Kary Mullis, 1944-
PCR Copies a Specific DNA Sequence
Fig. 13-3
1 2 3
1 2 4 8
PCR
cycles
DNA
copies
4
16
etc.
etc.
DNA
fragment
to be
amplified
original
DNA
194°F (90°C) 122°F (50°C) 158°F (70°C)
DNA
polymerase
new DNA
strandsprimers
Heating
separates
DNA strands
Cooling allows
primers and
DNA polymerase
to bind
New DNA
strands are
synthesized
1 2 3
(b)Each PCR cycle doubles the number of copies of the DNA(a) One PCR cycle
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Short-Tandem Repeats Are Common in Noncoding Regions of DNA
Fig. 13-4
Eight side-by-side (tandem) repeats
of the same four-nucleotide sequence
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STR #1: The probe base-pairs and binds
to the DNA
STR #2: The probe cannot base-pair with the DNA,
so it does not bind
probe
label
(colored molecule)
DNA Probes Base-Pair with Complementary DNA Sequences
Fig. 13-6
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DNA Profiling
Fig. 13-7
STR name
Penta D
CSF
D16
D7
D16: An STR on chromosome 16
DNA samples from
13 different people
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14
1312
11
10
98
Nu
mb
er
of
rep
eats
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Gel Electrophoresis and Labeling with DNA Probes Separates and Identifies
Segments of DNA
Fig. 13-5
nylon paper
solution of DNA probes
(red)
gel
power supply
wells
pipetter
DNA “bands”
(not yet visible)
gel
nylon paper
–+
–+
DNA samples are pipetted into wells
(shallow slots) in the gel. Electrical current
is sent through the gel (negative at the
end with the wells and positive at the
opposite end).
Electrical current moves the DNA
segments through the gel. Smaller
pieces of DNA move farther toward the
positive electrode.
The gel is placed on special nylon
“paper.” Electrical current drives the
DNA out of the gel onto the nylon.
Complementary DNA segments are
labeled by the probes (red bands).
The nylon paper with the DNA bound
to it is bathed in a solution of labeled
DNA probes (red) that are
complementary to specific DNA
segments in the original DNA sample.
1
2
3
4
5
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Electroforesis en geles de agarosa y poliacrilamida
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Electroforesis en geles de agarosa y poliacrilamida
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BIOTECNOLOGÍA FORENSE
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Pruebas de paternidad
Como el hijo tiene la mitad de sus alelos de la madre y la otra mitad del padre, en los perfiles electroforéticos
se debe ver dicha correspondencia y esto permite saber quién es el padre.
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Escenas de crimen
Comparando muestras obtenidas de la escena del crimen, la víctima y los sospechosos se puede determinar quién es el culpable.
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¿Culpable o inocente?
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BIOTECNOLOGIA VEGETAL
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Table 13-1
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Bt Plants Resist Insect Attack
Fig. 13-8
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Restriction Enzymes Cut DNA at Specific Nucleotide Sequences
Fig. 13-9
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Using Plasmids to Insert a Bacterial Gene into a Plant Chromosome
Fig. 13-10, step 1 of 4
DNA including
Bt gene (blue) Ti plasmid
The DNA containing the Bt gene and
the Ti plasmid are cut with the same
restriction enzyme.
1
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Using Plasmids to Insert a Bacterial Gene into a Plant
Chromosome
Fig. 13-10, step 2 of 4
recombinant Ti
plasmid with
Bt gene
Bt genes and Ti plasmids, both with the
same complementary sticky ends, are mixed
together; DNA ligase bonds the Bt genes into
the plasmids.
2
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Using Plasmids to Insert a Bacterial Gene into a Plant
Chromosome
Fig. 13-10, step 3 of 4
A. Tumefaciens
bacteriumbacterial
chromosomerecombinant Ti
plasmids
Bacteria are transformed with the
recombinant plasmids.
3
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Using Plasmids to Insert a Bacterial Gene into a Plant
Chromosome
Fig. 13-10, step 4 of 4
Bt gene
plant cellplant
chromosome
Transgenic bacteria enter the plant
cells, and Bt genes are inserted into the
chromosomes of the plant cells.
4
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Beneficios de los cultivos transgénicos
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BIOTECNOLOGÍA MÉDICA
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large piece
of DNAsmall piece
of DNA
(a) MstII cuts the normal globin allele into two
pieces that can be labeled by a probe
MstII
cut #1MstII
cut #3
MstII
cut #2
DNA probe
Diagnosing Sickle-Cell Anemia with Restriction Enzymes
Fig. 13-11a
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(b) MstII cuts the sickle-cell allele into one very
large piece that can be labeled by the same probe
very large piece
of DNA
MstII
cut #1MstII
cut #3
DNA probe
Diagnosing Sickle-Cell Anemia with Restriction Enzymes
Fig. 13-11b
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(c) Analysis of globin alleles by gel electrophoresis
larger pieces
of DNA
smaller pieces
of DNA
Homozygous
sickle-cell: one
band of very
large DNA
pieces
Homozygous normal:
one band of large DNA
pieces and one band
of small DNA pieces
Heterozygous:
three bands
Diagnosing Sickle-Cell Anemia with Restriction Enzymes
Fig. 13-11c
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A Cystic Fibrosis Diagnostic Array
Fig. 13-12
DNA probe
for normal
CFTR allele
DNA probes for
10 different mutant
CFTR alleles
(a) Linear array of probes for cystic fibrosis
(b) CFTR allele labeled with a colored molecule
#1
(c) Linear arrays with labeled DNA samples from three
different people
Homozygous for normal CFTR alleles—
the person is phenotypically normal
#2
One normal and one defective CFTR allele—
the person is phenotypically normal
#3
Two different defective CFTR alleles—
the person develops cystic fibrosis
colored molecule
piece of patient’s DNA
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A Human DNA Microarray
Fig. 13-13
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Table 13-2
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parents with a
genetic disease
fertilized egg with
a defective gene
embryo with a
genetic defect
therapeutic
gene
genetically corrected
cell from culture
egg cell without
a nucleus
genetically corrected
clone of the original
embryo
healthy baby
genetically corrected
egg cell
viral vector
cell removed
and cultured
treated culture
Using Biotechnology to Correct Defects in Human Embryos
Fig. 13-14
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