Applications of molecular techniques used in research

Batten Disease Neuronal ceroid lipofuscinosis (NCL)

Neurodegenerative lysosomal storage disorder

At least 13 distinct human NCL genes known, all causing the same clinical features of visual failure, seizures and progressive motor and cognitive decline

Naturally occurring animal models, including sheep, whose disease closely matches the human

No effective therapies

Batten Animal Research Network

Gene therapy virus

Disabled virus

Corrective NCL gene packaged into virus

In vitro (in the lab)

In vivo (in the animal)

Corrective protein produced

Vector inserts gene

Target brain cellNucleus

Gene therapy

Using DNA that encodes a functional, therapeutic NCL gene to replace the mutated or missing NCL gene

Injection of the viral vector containing the corrective NCL gene into the brain of affected sheep

Gene expression of the missing gene and production of the corrective protein

Corrective NCL gene

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Battens Disease - Research by Professor David Palmer

Transposons(Evolutions “dead-mans” switch)

• Mobile genetic elements = Transposons• Present in all organisms (yes - even Humans!!)

– The larger the genome the higher % of TEs present– Humans = 50% TEs– Grapes = 45% TEs– Wheat = >75% TEs

Transposons – things they do…..

• New insertions at new locations– Can destroy gene function– Can change gene activity– Can shuffle exons making new

genes with new functions

• Existing and new insertions can influence the function of DNA flanking genome insertion sites = one important aspect of epigenetics

Transposons – Parasites or an evolutionary force for good?

• Both! - They are parasites that have been harnessed by evolution

• Most TEs contain regulatory elements that respond to and are activated when the host plant is exposed to stress

• It appears that retention of TEs is an evolutionary adaptation to rapid environmental change

– Activation under stress conditions (release of the dead-mans switch) can introduce massive amounts of new genetic diversity within a single generation

Cyclodia

Transposon

Cyclodia methylation

Transposons & Functional genomics

Develop somatic embryo cultures

Activate TEs using stress

Recover plants

Genotype to identify new element insertions and their location

Regenerate plants

Screen data for mutation in key genes• Identify genes + mutation• Identify individuals in the population• Assess the phenotypic impact of the mutation• Determine the function of the gene

Screen population for new phenotypes• Identify new plants with desirable traits• Interrogate database to find out what

new mutations are present• Identify genes mutated• Use information to ascertain gene

function and /or use as markers for rapid breeding

Transposons –Research by Dr Chris Winefield

Powelliphanta augusta New species of endemic giant landsnail

Single population located within mine footprint

Snail collected for translocation and captive management

No idea of their feeding requirements

Snails are nocturnal and difficult to observe feeding

Molecular diet analysisUsing snail faeces

DNA barcoding on snail feces (non invasive method)

Detection of at least 18 species of earthworms eaten by the snails

Information useful for the selection of new translocation sites and to feed the snails in captivity

Discovery of new species of earthworms never found before

Snail speciation – Research by Dr Stephane Boyer