Bio unit 4

Types of Biotech

• selective breeding

• cloning plants

• genetically modified organisms

Molecular Cloning

1. amplify w/ PCR

2. cut out gene of interest

3. isolate DNA fragments of gene of interest on gel

4. put fragments into plasmid then into new organism

PCR

1. Denature/unwind DNA

2. primers anneal to template, no primase needed

3. polymerase elongates strands (taq polmerase)

DNA ligation

• cut vector and DNA w/ sticky ends

• fit vector and DNA fragment together

• ligase glues and seals fragments together

• vector goes into new organism (make sure it’s in nw population)

• Gene of Interest and the selection marker will live while everything else dies.

CRISPR/CAS9 (bacteria)

• 1st viral infection.

  • viral DNA fragments collected and incorporated into bacterial genome, viral DNA library called CRISPR

• 2nd viral infection

  • RNA from CRISPR library serves where to cut

  • Cas9 cuts DNA

  • re-infection prevented

CRISPR/Cas9 (gene editing)

• inject into cell

  • guide RNA, Cas9 ‘scissors’ and new DNA sequence

• DNA can be:

  • deleted, inserted, or single base pairs mutated (for curing illnesses)

Genetic Maps

show relative location of genes on a chromosome, more abstract

Physical maps

show actual locations of genes, uses restriction maps or cytological maps, shows nucleotide level of resolution

Restriction maps

diagrams that show the locations of restriction enzyme cut sites on DNA

cytological/chromosome maps

diagrams that represent the physical arrangement of genes on chromosomes, based on the staining patterns observed under a microscope.

somatic cells

• cannot make egg and sperm

• risks and benefits only impacts individual

germline cells

• can make egg and sperm

• offspring impacted and evolution impacts

Sequence Tagged Sites (STS)

HIGHEST resolution, molecular markers used in mapping genomes.

ddNTPS

once added in a sequence and that sequence will stop. They are chain-terminating nucleotides used in DNA sequencing. Have no oxygen present in their hydroxyl group.

Sequencing Steps

1. genome is fragmented and pieces are sequenced.

2. assembly. Fragments put in order with computer software. Shotgun or clone-contig method

3. Annotation. DNA sequence is labeled w/ information about different regions to create usable map.

Clone contig

breakdown into fragments, put clones back together using STS sites. Takes longer, slow and steady.

Shotgun method

whole genome is fragmented, use computer to put parts together. Typically done with a reference of a prior or target genome. Fast and dirty.

coding DNA

makes function RNA + proteins

• alternative splicing - what are exons, introns used for

• open reading frames

non-coding DNA

doesn’t make function RNA or proteins.

• where are introns

• presence of pseudogenes

• telomeres

comparative genomics

compares info from 1 genome to learn about 2nd genome

functional genomics

uses biotech to highlight connection btwn phenotype and genotype.

• study all RNA molecules produced by TRANSCRIPTOME

• study all proteins produced by PROTEOME

• study interactions between proteins

proteomics

study of proteome/collection of proteins encoded in the genome.

• mass spectrometry - change mass to no proteins/AA

• protein microarrays - rely on protein-specific antibodies to ID protein translated.

Differentiation

as stem cells lose potency, cell gains more functions

totipotent

fertilized egg, diploid. Baby + placenta

Pluripotent

form tissues of organism but not embryonic tissues (NOT placenta). Embryonic stem cells from blastocyst + inner cell mass

multipotent

only form muscle, nerve and adipocyte, adult cells.

Requirements for Development

• cell division

• differentiation

• pattern formation (blueprints)

• morphogenesis (building)

hox genes

highly conserved, where to lay blueprints, mutations can result in misID of embryonic segments or misplacement of body parts.

morphogenesis

have to replicate cells and give them shape. Utilizes cell builidng and cell death

Apoptosis

controlled cell death. Normal for development, healthy, dies cleanly, packaged + sent out, recycle parts.

Nuclear Transfer stem cells

• already differentiated cell

• injecting differentiated cell nuclues into egg donor, egg is then reinserted into original organism, making a clone!

• therapeutic cloning

induced pluripotent stem cells

• differentiated cell goes under transcription factors and comes out as a stem cell which is pluripotent.

• never involves a human blastocyst

• a little more articial

• has a tendency to revert back into differentiated cell

natural selection

• genetic variation must already exist prior to selection.

• selection must alter production of offspring

• trait must be inherited

• sexual selection!

Gene flow

migration, babies pass onto the next generation.

Genetic drift

chance. Big population less effected than small population.

bottleneck effect

involves death, whatever left will be passed on

founder effect

small group moves on to another place, no death, live <3

sympatric

share same space, different parts (Think APES birds)

allopatric

live in different places divided by geographical barrier

ecological isolation

pre zygotic, same space though organisms are not interacting

behavioral isolation

pre zygotic, i see you though im not interested!

temporal isolation

pre zygotic, time (dawn v dusk)

mechanical isolation

pre zygotic, the parts just don’t fit

prevention of gamete fusion

pre zygotic, sperm and egg just aren’t compatible

hybrid invaibility

post-zygotic, unable to have live birth

hybrid infertility

post-zygotic, birth of offspring but offspring cannot reproduce

biological species concept

species is a group that breeds/could breed togther.

doesn’t reproduce w/ anyone else and if they did, that offspring would be infertile.

only works on live organisms

phylogenetic species concept

population that has been evolving independently from other groups.

identify through phylogenetic analysis

overestimate species

reinforcement

process where natural selection strengthens reproductive isolation between diverging species

adaptive radiation

not evolving one at a time, instead evolving all at once

gradualism

relatively slow rate of mutations, long + slow

punctuated equilibrium

periods of faster rates of mutations, could include adaptive radiation!

node

point of divergence in a phylogenic tree

synapmorphy

characteristic/trait shared by group of organisms and believed to originate in most recent common ancestor. looks same, same ancestor (lactation)

ancestral trait

similar traits PRIOR to common ancestor

derived trait

similarity between species inherited from most recent common ancestor

systematics

study of evolutionary relationships and reconstruction

outgroup

closely related but not direct members being studied

clade

species that share common ancestor

parismony

theory that requires the fewest evolutionary steps is considered the best hypothesis

homoplasy

shared characteristic state, not inherited from common ancestor, can result from convergent evolution or evolutionary reversal.

animals look the same but have different ancestors (bats and birds)

monophyletic

whole clade

paraphyletic

almost everything but excludes a node

polyphyletic

cherrypicking

homology

assume farther apart than they actually are, looks different but has same ancestor (hands and hooves and bat wings)

cladistics approach

uses parsimony, simplest answer and add up synapomorphies

molecular clock

add time data,, assumes steady change over time

statistics approach

punctuated equilibrium, adaptive radiations, parsimony doesn’t apply, computers needed

Hadean Eon

lots of volcanoes, lightening, no life, organic compounds where proven bc of miller urey experiment.

Archean Eon

2nd eon, life starts, its an RNA world (DNA ← RNA → proteins)

Proterozoic eon

• mitochondria allow for multicellularity

• sexual reproduction

• extra membranes

• endosymbiosis

phanerozoic eon

• cambrian explosion!

• world gets used to oxygen

  • skeletal arms race (great fossils!)

Radio Isotope dating

• add numbers for total atoms

• divide first number by total atoms

• find that percentage on y-axis

• follow on half-life line

• follow down to find number of years