Development and Biological Tools

Natural selection has favored alleles that increase…

SA, leading to features like branched alveoli

John Gurdon Experiment

Replaced egg nucleus with skin cell nucleus and skin cell DNA can still function

If all cells have the entire genome, why do different cells express different proteins?

Alternative splicing, Different parts of genome activated and repressed, different TF being active, Translational regulation, Translational degradation

Translational Regulation

Blocking ribosomes from binding, using miRNAs, Alternating mRNA structure so ribosome can’t bind, controlling initiation factors

Translational degradation

Newly synthesized proteins or corresponding mRNA’s are broken down while translation is still occuring

What do model organisms need to have/do

Grow easily, easy to study (able to genetically modify, sequenced genome), quickly reproducing, research community

Examples of model organisms

mouse, fruit fly, yeast, arabidopsis

Mutagenesis Screens

Take plants and induce lots of mutations using (UV, EMS which induces a bunch of substitiutions), results: lots of plants with lots of mutations —→ look at phenotypes, mutations that affect plant development

How do I know which gene mutations are in

Next Gen Sequencing (PCR), we need to identify the specific mutation that is causing the messed up phenotype (pick candidate genes and rank genes based on what it involves)

Expression Pattern

Is the gene turned on in the right organs at the right place and right time

If the gene is necessary for the process..

Outcome will not occur without factor

If the gene is sufficient

Factor will ALWAYS cause outcome

In Situ Hybridization

Fluorescently label RNA/DNA complementary to the mRNA of target gene, add it to cells, see mRNA expression (only happens in dead tissues (have to open the cell to immobilize)

When do we use in situ hybridization

To find out where mRNA is, or if regulation is happening via degradation of protein

Fluorescent antibody staining

Antibodies (immune system proteins that can be easily developed to bind to any specific protein) with fluorescent markers against target gene, add them to cells, look for protein expression

3 ways to test expression patterns

fluorescent protein expression

Genes encoding fluorescent proteins can be fused to the gene of interest (or regulatory DNA) to see protein expression

GFP

Wherever gene is expressed so if GFP (fusion of protein + GFP)

AP3

Part of a group of genes that tells us how flowers form different parts in specific positions

AAP3 is expressed in

the ring of cells in the developing flower (Helps plants make petals and stamens

What happens if we remove AP3

No petals form necessary for development

CRISPR

Can cut DNA at any sequence guided by 20 bp of guide RNA

How does the CRISPR method work

After cut, the mechanism to repair the DNA would ligate the broken ends together without a template, errors occur and the repair can add/remove bases causing a frameshift mutation that disrupts the gene

Overexpression

Forcing a gene to be expressed more than usual in places where its not normally active. Done by inserting a copy of the gene under a strong promoter in plants genome, adding regulatory DNA

By forcing AP3 into new areas we can see which

organs can be converted into petals/stamens

If we add AP3 gene with AP3 regulatory DNA, where will it be expressed?

Its expression is controlled by its regulatory DNA so the gene will be expressed in the same domain

PAX6

Transcription factors responsible for initiating eye development across many animals

Overexpression of PAX6 results in

Formation of whole eyes in wrong places

3 classes of genes responsible for the spatial pattern of floral parts

A, B, C regulate expression of other genes responsible for sepals, petals, stamens, and carpals

Combinatorial Control

TF work together to regulate expression of other genes, by combining in different ways, a small number of factors can regulate a large number of genes

Cells can integrate multiple signals through turning on

Combinations of TF

Stem Cells

They are able to make more cells like themselves (can self renew) and they can become other cells that have differen functions (differentiation), found in almost all tissues in the body

Stem cells are needed for

maintenence of tissues as well as repair after injury

if the body needs more stem cells it will divide in a specific way to

result in 2 stem cells (usually during development/injury)

Different ways stem cells can divide

• 1 stem cell → 1 stem cell + 1 differentiated cell (increases differentiation)

• 1 stem cell → 2 differentiated cells (reduces the stem cell pool and increases differentiated cells quickly (used when tissues needs lots of mature cells fast

Anatomy

study of the biological form of an organism

Physiology

Study of the biological functions an organism performs

Proximate level of causation

Researchers who focus on the physical/chemical mechanisms responsible for function

Ultimate Level of Causation

Researchers who focus on how physiological systems evolved and how they impact fitness