טכניון מכון טכנולוגי לישראל
הטכניון מכון טכנולוגי לישראל - בית הספר ללימודי מוסמכים  
Ph.D Thesis
Ph.D StudentRegev Issachar
SubjectGenetic Approaches for Gene Tagging and Isolation
of Functional Genes in Triploid Banana Plants as
Means for Biotechnological Applications
DepartmentDepartment of Biology
Supervisors Professor Emeritus Shimon Gepstein
Dr. Eli Khayat


Abstract

Given that banana and plantain comprise the  major source of energy in the diet of well over 400 million people around the world, it was ranked by the FAO the fourth important food-crop after wheat, rice and maize. Most edible bananas are triploid, seedless, and reproduce vegetatively. The majority of producers and consumers of banana and plantains are under-developed countries that have neither technical means, nor budgets for carrying sophisticated breeding programs. The complexity of breeding triploid and infertile plant species is a major constrain to breeders who are mainly seeking disease resistance and high fruit quality traits. The main objective of the current research was to establish an infrastructure for biotechnological applications in banana germplasm improvement. In order to achieve this goal, two strategies were employed:

  • Implementing transposon mutagenesis as a tool for gene tagging in the Musa genome.  To this end, the maize Ac transposable element system was introduced to bananas.
  • Identification, characterization and isolation of genes specifically related to fruit ripening and senescence. This will allow future manipulation of these genes.

Using particle bombardment of an embriogenic cell suspension culture, we have generated transgenic banana lines, harboring the maize Ac transposable element. Both excision and re-insertion of the Ac element were demonstrated. In order to stabilize  the Ac element in the transgenic plants (not by using cross hybridization), we have  assembled a construct containing antisense RNA for the transposase segment of the open reading frame. This construct also showed excision activity.

We have used the suppressive subtractive hybridization method for the isolation of ripening and senescence related cDNA clones.  Several isolated clones have related homologues in public genebank data bases and are related to fruit ripening or senescing tissues, while the sequence analysis of others did not show resemblance to annotated genes. Ethylene treatment of harvested fruit induced expression of part of the genes, while other clones were unaffected. The expression pattern of part of the clones was examined in several genotypes of banana and plantains in the course of ripening and senescing processes and under ethylene treatments of the fruit. A distinctive ripening pattern was exhibited by the different banana and plantain genotypes.