Oregon State University

Transgenics

Transgenic Plants

I. Why create transgenic plants?

When there is no naturally occurring genetic variation for the target trait. 

Examples: 

  1. Herbicide resistance
  2. Vitamin A
  3. Flower color
  • Florigene. L-3. A company dedicated to novel flower colors, through transgenics 

 

IIWhat genes to transfer?

  1. One gene to a few genes - the CP4 ESPS example
  2. Multiple genes - the “Golden Rice” example
  3. In principle, any gene, 
  • CP4 ESPS L-2. Details on the gene conferring resistance to the herbicide RoundUp. The gene conferring resistance to Roundup herbicide was found in Agrobacterium. Coincidentally, this organism is also a tool for creating tansgenic plants
  • "Golden Rice”. L-1. A biosynthetic pathway with genes from daffodil, bacteria, and pieces of genes from pea, rice, and virus.

III. Constructing transgenes. The minimal requirements for the transgene are a promoter, coding region, and terminator.

General structure: 5’---Promoter …..Coding region…….terminator---3’

Example (Bt gene with nptII selectable marker): 5’ --P35S…Bt…Tnos--//--P35S…nptII…Tnos---3’

A. Promoter: DNA sequence controlling spatial and/or temporal level of transgene expression. Promoters can be

  1. constitutive (e.g. CaMV 35S, Ubiquitin, NOS = nopaline synthase)
  2. tissue specific (e.g. patatin , which is tuber specific)
  3. inducible (e.g. heat shock, photo responsive)

B. Termination sequence:  DNA sequence signaling end of the gene (e.g. T35S, T NOS).

In addition to the target gene, additional elements may be added, such as selectable markers and/or reporter genes

C. Selectable Marker:  Encodes a protein (enzyme) that allows the transformed cells to grow while the growth of the non-transformed cells is inhibited. Examples include

  1. Antibiotic resistance (e.g. nptII, hygromycin phosphotransferase)
  2. Herbicide resistance (e.g. bar, phosphinothricin acetyl transferase)
  • Selectable Markers. L-1. Key types of selectable markers and L-3. Details and examples of selectable markers.

D. Reporter genes:  Genes that, upon expression in the transgenic plants, provide a clear indication that genetic transformation did occur, and indicate the location and the level of expression. Examples include. 

 

  1. Glucuronidase (GUS) - open source molecular biology tool arising from the discovery of GUS - CAMBIA 
  2. Luciferase, green fluorescent protein (GFP). An extreme use of GFP
GUS. L-2. Blue means GUS is expressed.
CAMBIA. L-3. Royalty income from GUS used to establish a biotech non-profit.
GFP. L-3. So many ways to be green.
An extreme use of GFP. L-3. Art meets science (?). 

 

E. Additional components which may be included in the transgene are enhancers and introns.

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