Gene Design Flashcards
dicot or monocot
tobacco
Dicot
dicot or monocot
Cotton
Dicot
dicot or monocot
Soybean
Dicot
dicot or monocot
Legumes
Dicot
dicot or monocot
Arabidopsis
Dicot
dicot or monocot
Wheat
monocot
dicot or monocot
corn
monocot
dicot or monocot
Rice
monocot
dicot or monocot
Barley
dicot or monocot
how does agrobacterium mediated transfer work
Ti plasmid to allow transfer of T-DNA section containing transgene to plant, bacteria incubated with leaf discs
Cry1
Lepidoptera
Cry3
Coleoptera
Cry2
Diptera
What species produces Cry toxins
Bacillus thuringiensis
When was Bt a registered pesticide
1961
Cry toxins have a what structure?
What are the domains?
Conserved structure 600aa
3 distinct domains
1-N terminus 7alpha helix membrane insertion pore formation
2- B prism, receptor recognition and binding
3- C terminal antiparallel B sheets receptor recognition and pore formation
Digestion of Cry toxins does what
Activates then
Digestion to N terminal 60-70kDa truncated form
“Toxin forms lytic pores in cell membrane of insect guy and once digested (active) indigested (inactive)
The toxin binds where?
This process is called ?
To the high affinity receptors in midgut epithelial
- forms pores
- kills e cell
Colliod osmotic lysis
What is a protoxin
Proteolyticall cleaved/inbound to receptor = not toxic “protoxin”
NOT SURE TBH Might he wrong
Lepidoptera
Butterflies and moths
Diptera
Flies and mosquitoes
Coleoptera
Beetles and weevils
Hymenoptera
Wasps and bees
Herbicide res Maize
Round up
Maize = monocot
P - Ubi
FG - aroA (glyphosate red, detoxifies)
T - Nos (nopaline synthase)
MG - nptII kanomycin res
Says for cereals such a maize better to use herbicide selection gene - eg Hydromycin bar gene
Aluminium tolerant legumes
P- CamV35s
FG- Csb (citrate synthase gene from pseudomonas) - citrate increases too by chelating Al3+
T- NOS nopaline synthase terminator
M- NptII neomycin
Gene construct in cytoplasm not mitochondria so doesn’t interfere with Krebs cycle
Butterfly insecticide res in rice
Rice = monocot
P -Ubi
FG - Cry1 from bt
T - Nos nopaline synthase term
P-Ubi
FG- CPT1A (protease inhibitor)
T-Nos
P-Ubi
MG- nptII rice :( kanomycin therefore use paromomycin
T-Nos
Herbicide too cotton
Cotton = Dicot
P-CamV35S
FG - bar (against glufosinate)
T- nos
P-CamV35S
MG- nptII kanomycin
T-nos
Tobacco drought/salinity tol
P- SARK (senescence associated receptor protein kinase) promoter
TG- IPT (isopentyl transferase, cytokinin biosyn, prevent senescence)
T-Nos nopaline syn term
P-CamV35s
MG- nptII
T-Nos
Soybean drought tol
P-CamV35s
FG- AtZEP (zeaxanthin epoxidase gene makes zeaxanthin precursor to ABA)
T- Nos
P-CamV35s
FG-nptII kanomycin
T-nos
Example of light inducible promoter
Promoter from cab gene
Use what if don’t what expression all over the plant?
Signal transit peptides
Can be used to direct the gene construct to the desired part of the plant
E.g. chloroplast or endosperm
Endosperm useful in rice as a lot of metabolic activity takes place here
Drought tol maize
P-Ubi
TG- AVP1 (arabidopsis H+-Ppase pump)
T-nos
P-Ubi
MG- bar (herbicide selection w/ PPT)
what is a newly developed stacking system
Zhu et al (2017) have developed a transgene stacking vector system (TGSII) which allows the successful transformation of plants (tested in rice) with several genes in one construct, each with their own promotors and terminators.
How does the TGSII stacking system work
These utilise antibiotic selection cassettes (nptII) and Cre/loxp-mediated constructs which allows the marker cassette to be removed under tissue-specific promotors, once the transformants have been selected. This allows a marker free gene construct in planta and it also works with agrobacterium transformation. This could be an interesting alternative for a company to try, as it has had proven success with crop such as Astaxanthin bio-engineered rice (Zhu et al. 2018).
