Methods for large scale functional evaluation of nucleotide sequences in plants

Abstract

The present invention provides for rapid and large scale evaluation of expression of, or function of, nucleotide sequences in plants. The invention comprises three specific components which provide for fast and large scale evaluation of nucleotide sequences. The first component includes delivery in either a single event a library of different engineered vectors or a single engineered vector for a single target nucleotide sequence comprising sequences the function of which is desired to be known in plant cells. Surprisingly, applicants have discovered that, the introduction of multiple vectors to plant cells predominantly results in individual transgenic plants which contain only a single transformation event. The second feature of the invention involves a highly transformable, fast cycling and/or miniature size plant and the final step involves mass scale analysis of T0 plants for various phenotypes and plasmid rescue to identify the nucleotide sequence present in a particular phenotype.

1 .- 22 . (canceled) 
     
     
         23 . A method of analyzing plants having a phenotype of interest comprising:
 obtaining a population of plants grown under greenhouse conditions,   obtaining phenotypic characteristics of the population of plants,   comparing phenotypic characteristics of plants in the population to one another to   identify phenotypic differences among the plants,   identifying plants having a phenotype of interest, and   selecting plants with said phenotype of interest.   
     
     
         24 . The method of  claim 23 , wherein the phenotypic characteristics are recorded. 
     
     
         25 . The method of  claim 23 , wherein the identified phenotype is associated with the specific plant having said phenotype. 
     
     
         26 . The method of  claim 23 , wherein the phenotypic characteristics of the plants are recorded and associated with the identity of the particular plant having the recorded phenotype. 
     
     
         27 . The method of  claim 23 , wherein the compared phenotypic characteristic is stalk strength, height, seed set, maturity, leaf necrosis, leaf senescence, or tassel branch number. 
     
     
         28 . A high throughput method of assessing a plant trait of interest comprising:
 providing a population of plants with different genotypes,   growing the plant population under greenhouse conditions,   analyzing individual plants having the trait,   comparing plants having the trait to other plants in the provided population, to identify the genetic basis of the trait.   
     
     
         29 . The method of  claim 28 , wherein the individual plants are visually analyzed. 
     
     
         30 . The method of  claim 28 , wherein an observed phenotypic characteristic is associated with the specific plant having said trait. 
     
     
         31 . The method of  claim 28 , wherein the phenotypic aspects of the analyzed plants are recorded and associated with the identity of the particular plant having the recorded phenotype. 
     
     
         32 . The method of  claim 28 , wherein the phenotypic characteristic analyzed is stalk strength, height, seed set, maturity, leaf necrosis, leaf senescence, or tassel branch number. 
     
     
         33 . A method of selecting plants having a phenotype of interest comprising:
 obtaining a population of plants grown under greenhouse conditions,   visually screening individual plants in the plant population to identify phenotypic characteristics,   comparing plants in the screened population to one another to identify phenotypic differences among the plants,   identifying plants having a phenotype of interest, and   selecting plants with said phenotype of interest.   
     
     
         34 . The method of  claim 33 , wherein the results of visual screening are recorded. 
     
     
         35 . The method of  claim 33 , wherein an observed phenotypic characteristic is associated with the specific plant having said phenotypic characteristic. 
     
     
         36 . The method of  claim 33 , wherein the phenotypic aspects of the screened plants are recorded and associated with the identity of the particular plant having the recorded phenotype. 
     
     
         37 . The method of  claim 33 , wherein the phenotypic characteristic analyzed is stalk strength, height, seed set, maturity, leaf necrosis, leaf senescence, or tassel branch number. 
     
     
         38 . A high throughput method of assessing a plant trait of interest comprising:
 obtaining a population of plants with different genetic constitutions,   growing the plant population under greenhouse conditions,   visually screening individual plants having the trait,   comparing plants having the trait to other plants in the obtained population, to identify the genetic basis of the trait.   
     
     
         39 . The method of  claim 38 , wherein the results of visual screening are recorded. 
     
     
         40 . The method of  claim 38 , wherein an observed phenotypic characteristic is associated with the specific plant having said phenotypic characteristic. 
     
     
         41 . The method of  claim 38 , wherein the phenotypic aspects of the screened plants are recorded and associated with the identity of the particular plant having the recorded phenotype. 
     
     
         42 . The method of  claim 38 , wherein the trait analyzed relates to stalk strength, height, seed set, maturity, leaf necrosis, leaf senescence, or tassel branch number. 
     
     
         43 . A method of analyzing plants expressing a target gene the method comprising:
 obtaining a population of plants grown under greenhouse conditions,   obtaining phenotypic characteristics of the population of plants,   comparing phenotypic characteristics of the plants,   analyzing phenotypic differences among the plants; and   identifying plants that are relevant to the trait of interest.   
     
     
         44 . The method of  claim 43 , wherein the population of plants comprises corn. 
     
     
         45 . The method of  claim 44 , wherein the corn is a fast cycling corn line. 
     
     
         46 . The method of  claim 45 , wherein the fast cycling corn line is Gaspe Flint. 
     
     
         47 . A method of analyzing transgenic plants expressing a trait of interest:
 obtaining plant cells for transformation,   obtaining a plurality of nucleotide vectors,   introducing the plurality of vectors to form a transgenic event population comprising transgenic plants,   growing the transgenic plants under greenhouse conditions,   visually screening individual plants having the trait; and   identifying plants that are relevant to the trait of interest.   
     
     
         48 . The method of  claim 47 , wherein the trait is stress response. 
     
     
         49 . The method of  claim 47 , wherein the transgenic plants comprise corn. 
     
     
         50 . The method of  claim 49 , wherein the corn is a fast cycling corn line. 
     
     
         51 . The method of  claim 49 , wherein the fast cycling corn line is Gaspe Flint.