Berthold’s Experiment: Introduction

• Background: pangenesis/preformation theory of reproduction– Idea that various body parts secreted bits of

themselves into the blood and these assembled in semen. Preformation extends this to teeny animals (homunculi) being formed in sperm in the testes. These homunculi then simply grew larger to form babies.

• Not a very promising start, but it had some truth that helped Berthold:– Body parts able to secrete factors into blood– These factors have targets distant to origin– The testes were somehow involved in this

Berthold’s Hypothesis

• According to pangenesis/preformation, bits of organs travel through the blood stream to the testes to make a copy of that animal. This theory makes a specific prediction that seems testable:

Hypothesis: transplanted testes should pass on the properties of host and not the donor to offspring…

(N.B. this is my guess as to why Berthold did his experiment, we don’t really know why)

Berthold’s Experiment: Methods

• Experimental design:– control group 1: castrated only

Concern: What are effects of castration on roosters?– control group 2: castrated and testes reimplanted

Concern: Will testes be functional after removal and implantation?

– experimental group: castrated and cross- implanted

Will testes function like the host (support pangenesis) or like the donor (refute pangenesis).

Unfortunately for our imagined hypothesis, implanted roosters aren’t fertile. So we get no further towards disproving pangenesis.

BUT…serendipity!

TARGET PROBE LABEL IN SITU IN VITRO

Hormone

(non-protein)

Antibody Isotope

Enzyme

Not Common

RIA

EIA/ELISA

Enzyme Assays

Protein Antibody Isotope

Enzyme

Fluorophore

IHC

(not ICC!)

Western

RNA cDNA Isotope

Enzyme

Fluorophore

ISH Northern

RT-PCR

microarray

DNA cDNA Isotope

Enzyme

Fluorophore

Not common

Southern

PCR

RFLP

Microarrays

• Relatively new technology. • Different kinds, idea is that the expression of

1000s of genes can be determined at once using an array of very small dots, each of a specific cDNA

• This kind of “high throughput” sampling of gene expression is very fashionable

Benefit: lots of information fastCost: expensive, validation and analysis is

laborious, often inconclusive (fishing expedition)

Genetically modified organisms

• Transgenesis (designer genes)Artificial genes that are added to

genome• Gene Targeting (knock-outs/ knock-

ins)Artificial genes that replace genomic

sequencesBenefit: very powerful way of testing

gene functionCost: laborious & time intensive,

abnormal genetic makeup often complicates study

Temporary genetic manipulations• Antibody binding can be used to interfere with

protein function as well as for detection• RNAi = RNA interference (a.k.a. antisense). cRNA

or cDNA can reduce protein abundance of specific genes

• Transfection (a.k.a. gene therapy). Introduction of foreign genes into cells using vectors (eg viruses)

Benefits: less time-intensive than GMOs, can also overcome developmental and compensatory effects often seen in GMOs

Costs: transient, variable, inefficient