Development of Crop Plants - History Domestication of crops We have been modifying our crops for 10,000 years through Selection. All crops we grow today have undergone extensive genetic change from their wild ancestors. Crops, strains and genes have moved around the globe. Domestication of corn creating a new species in the process – Zea mays Domestication of corn creating a new species in the process – Zea mays Thanks to C.S. Prakash for some of the pictures on this and the next couple of slides.

The domestication of animals has also been accompanied by extensive human-directed genetic modification. This is exemplified with the domestic dog, Canis familiaris. Modern genetics including genomics indicates that all domestic dogs are originally derived from the wolf, Canis lupus. Thus the Siberian Husky and Mexican Chihuahua are derived from the same original specie with the only difference being the extent of the genetic modification!

Traditional Breeding Crossing individuals with desirable characteristics (e.g. yield) and selecting among the progeny. Genes recombine in a random fashion and finding superior progeny has been as much art as science. Needs excellent management since it’s a numbers game and genotype often masked by environmental influences. Traditional Breeding Crossing individuals with desirable characteristics (e.g. yield) and selecting among the progeny. Genes recombine in a random fashion and finding superior progeny has been as much art as science. Needs excellent management since it’s a numbers game and genotype often masked by environmental influences. For specific characteristics such as fatty acid % of oil, naturally occurring mutants screened for and if not found induced by chemical mutagenesis or irradiation.

Conventional: Wide Hybridization introduces 20,000 to 100,000 potentially negative genes in order to obtain one desirable disease resistance gene. Induced mutagenesis has been used for decades to create genetic variants.

Genetic Engineering: Introduce one (or a few) foreign “good” genes into the best accepted cultivar background.

Genetic Engineering: Genetic Engineering: Introduce one (or a few) foreign “good” genes into the best accepted cultivar background. The main thing that is new with genetic engineering is that species barriers can now readily be bridged.

Genetic Engineering: Genetic Engineering: Introduce one (or a few) foreign “good” genes into the best accepted cultivar background. The main thing that is new with genetic engineering is that species barriers can now readily be bridged. This opens new opportunities and depending on how it is used requires new safeguards.

Recombinant DNA?

DNA_Extraction.swf

Gene Cloning

Agrobacterium tumefaciens A. tumefaciens Chromosome T-DNA Ti plasmid T-DNA Chromosomal DNA Crown Gall Plant crown gall tissue naturally transgenic Plant chromosomes containing T-DNA

How Do You Introduce a Foreign Gene into a Recipient Organism ? Overview

Methods Used to Date for Plant Transformation Agrobacterium tumefaciens and rhizogenes Gene Gun Microprojectiles (PDS) Electroporation of protoplasts Microinjection Pollen Tube Pathway Silica Carbide fibers Microlaser Viruses [some native genes replaced]

plasmid Desired DNA Recombinant plasmid Agrobacterium tumefaciens containing Wt Ti plasmid A. tumefaciens containing engineered Ti plasmid Plant cell inoculated with A. tumefaciens Plant cell containing Desired DNA Cultured plant cells Regenerant Adult plant expressing desired trait (DNA) Inserting foreign genes into plant cells. A plasmid containing DNA is cut with a restriction enzyme & DNA of desired gene (red) inserted. Desired gene then inserted into Ti (tumor-inducing) plasmid naturally found in A. tumefaciens. Plant cell inoculated with A. tumefaciens containing engineered Ti plasmid + the desired DNA transfers desired DNA + t-DNA into plant chromosomes. Plantlets with desired trait then regenerated.

The Gene Gun PDS1000 Microparticle Delivery System Helium chamber Rupture disk Macrocarrier DNA coated gold particle Stopping screen Focusing device Target tissue Gene gun From Collins lab

Biolistic Transformation Before impact DNA coated gold particle After impact During impact ? Plant Cell Wall

How do Genes do Their Job?

GENE EXPRESSION DNA Transcription mRNA Translation Protein

Product AProduct B Structural Gene Enzyme (Protein)

Roundup Ready Crops Monsanto web page:

Roundup (Glyphosate) is a very strong inhibitor of EPSP 1 Synthase. Glyphosate Shikimate 3-phosphate EPSP PEP 1 EPSP = 5-ENOLPYRUVYLSHIKIMATE 3-PHOSPHATE

Sulfonylurea Tolerant Soybeans - STS - These cultivars are resistant to certain sulfonyl ureas (SUs), a family of herbicides which are most effective against broadleaf weeds. STS herbicides used over soybean varieties that have the STS gene offer the benefit of using broad spectrum sulfonylurea broadleaf herbicides without injuring young soybean plants. Labeled sulfonylurea herbicides include Synchrony STS¹, Reliance STS¹, Classic¹, Pinnacle¹, Canopy¹, Canopy XL¹ and Concert¹. The STS gene was incorporated into soybean germplasm using conventional breeding methods. The STS gene was incorporated into soybean germplasm using conventional breeding methods. SU tolerant gene induced by EMS mutagenesis (US patent # 5,084,082). Bx breeding

SUs inhibit the essential plant enzyme acetolactate synthase or ALS. Animals do not have ALS. Mechanism of action of SUs  -keto butyrate pyruvate acetolactate ALS isoleucine CO 2

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Charles Darwin... “ I t is not the strongest species that survive, nor the most intelligent, but the ones most responsive to change” “I’m all for progress; it’s change I don’t like” - Mark Twain Slide from C.S. Prakash