Tomatoes That Soften More Slowly Post-Harvest Due To Non-Transgenic Alterations In An Expansin Gene

Abstract

A series of independent human-induced non-transgenic mutations found in an expansin gene (LeExp1) of tomato; tomato plants having these mutations in their LeExp1 genes; and a method of creating and identifying similar and/or additional mutations in the LeExp1 gene by screening pooled and/or individual tomato plants. The tomato plants of the present invention exhibit fruit that soften more slowly post-harvest without having the inclusion of foreign nucleic acids in their genomes.

1 . A method of producing a tomato plant with a human-induced non-transgenic mutation in an expansin gene having substantial homology to SEQ ID NO: 6 comprising the steps of:
 a. obtaining plant material from a parent tomato plant;   b. treating said plant material with a mutagen to create mutagenized plant material;   c. analyzing said mutagenized plant material to identify a plant having at least one mutation in at least one expansin gene having substantial homology to SEQ ID NO: 6.   
     
     
         2 . The method of  claim 1  wherein the mutation is selected from the group consisting of G220T, G274A, C305T, G403A, C460T, G937A, G940T, C986T, A991G, and G1001A 
     
     
         3 . The method of  claim 1  wherein the plant material is selected from the group consisting of seeds, pollen, plant cells, or plant tissue. 
     
     
         4 . The method of  claim 1  wherein the mutagen is ethyl methanesulfonate. 
     
     
         5 . The method of  claim 4  wherein the concentration of ethyl methanesulfonate used is from about 0.4 to about 1.2%. 
     
     
         6 . The method of  claim 1  further comprising the steps of:
 a. isolating genomic DNA from the mutagenized plant material or a progeny tomato plant; and 
 b. amplifying the isolated genomic DNA using at least one primer specific to an expansin gene having substantial homology to SEQ ID NO: 6. 
 
     
     
         7 . The method of  claim 1  further comprising the steps of:
 a. isolating genomic DNA from the mutagenized plant material or a progeny tomato plant; and 
 b. amplifying the isolated genomic DNA using at least one primer specific to DNA sequences adjacent to an expansin gene having substantial homology to SEQ ID NO: 6. 
 
     
     
         8 . Tomato fruit, seeds, pollen, plant parts or progeny of the tomato plant of  claim 1 . 
     
     
         9 . The tomato fruit of  claim 8 , wherein the tomato fruit have a decreased rate of post-harvest softening caused by a human-induced non-transgenic mutation in at least one LeExp1 gene. 
     
     
         10 . Food and food products incorporating the fruit of  claim 9 . 
     
     
         11 . The method of  claim 1  wherein the at least one mutation in at least one expansin gene codes for a protein comprising the mutation G65* in SEQ ID NO: 11. 
     
     
         12 . The method of  claim 1  wherein the at least one mutation in at least one expansin gene codes for a protein comprising the mutation G83R in SEQ ID NO: 11. 
     
     
         13 . The method of  claim 1  wherein the at least one mutation in at least one expansin gene codes for a protein comprising the mutation T93I in SEQ ID NO: 11. 
     
     
         14 . The method of  claim 1  wherein the at least one mutation in at least one expansin gene codes for a protein comprising the mutation G126S in SEQ ID NO: 11. 
     
     
         15 . The method of  claim 1  wherein the at least one mutation in at least one expansin gene codes for a protein comprising the mutation L145F in SEQ ID NO: 11. 
     
     
         16 . The method of  claim 1  wherein the at least one mutation in at least one expansin gene codes for a protein comprising the mutation G190R in SEQ ID NO: 11. 
     
     
         17 . The method of  claim 1  wherein the at least one mutation in at least one expansin gene codes for a protein comprising the mutation D191Y in SEQ ID NO: 11. 
     
     
         18 . The method of  claim 1  wherein the at least one mutation in at least one expansin gene codes for a protein comprising the mutation P206L in SEQ ID NO: 11. 
     
     
         19 . The method of  claim 1  wherein the at least one mutation in at least one expansin gene codes for a protein comprising the mutation 5208G in SEQ ID NO: 11. 
     
     
         20 . The method of  claim 1  wherein the at least one mutation in at least one expansin gene codes for a protein comprising the mutation W211* in SEQ ID NO: 11. 
     
     
         21 . An endogenous expansin gene having substantial homology to SEQ. ID NO: 6 and having at least one human-induced non-transgenic mutation within said endogenous expansin gene, wherein said at least one human-induced non-transgenic mutation is selected from the group consisting of G220T, G274A, C305T, G403A, C460T, G937A, G940T, C986T, A991G, and G1001A. 
     
     
         22 . A tomato plant containing the endogenous expansin gene of  claim 21 . 
     
     
         23 . Tomato fruit, seeds, pollen, plant parts, and progeny of the tomato plant of  claim 22 . 
     
     
         24 . The tomato fruit of  claim 23 , wherein the tomato fruit have a decreased rate of post-harvest softening caused by a human induced non-transgenic mutation in at least one LeExp1 gene. 
     
     
         25 . Food and food products incorporating the fruit of the tomato plant of  claim 24 . 
     
     
         26 . The endogenous expansin gene of  claim 21  wherein said human-induced non-transgenic mutation creates an amino acid change in the expansin protein expressed from the expansin gene, wherein said amino acid change is selected from G65*, G83R, T93I, G126S, L145F, G190R, D191Y, P206L. S208G, W211*. 
     
     
         27 . An expansin protein expressed from the endogenous expansin gene of  claim 26 .