Map Based Cloning System in Madicago Suitable for Isolating Genes Involved in Leaf Morphogenesis, Nodule Formation and Effectiveness of Nitrogen Fixation

In higher eukaryotic plants, the number of the genes is approximately 50, 000, therefore the plant genomes are fairly complex, consequently it is very hard to correlate genes with functions of an appropriate biological process, like symbiotic nitrogen fixation. Some techniques which are applicable for prokaryotic gene isolation (e.g. complementation of mutants) can not be used for most of the plant species. In order to correlate functions with cloned genes indirect biochemical approaches have been developed, like differential hybridization, construction of subtraction libraries, etc. Another laborious biochemical possibility is to identify a gene by its protein product. This approach involves the identification, purification and sequencing of a protein followed by the synthesis of specific oligonucleotides to use as hybridization probe or as primers for PCR. Fortunately there is an additional possibility by which specific genes can be identified: this is the genetic approach. In this case a mutant plant impaired in the appropriate biological process has to be isolated, then the gene suffered the mutation has to be genetically mapped using molecular markers, and finally the gene has to be cloned with the help of tightly linked molecular markers. This approach is the so-called map-based cloning strategy. Though the genetic strategy can not be used for every cases, nevertheless in those ones where applicable it provides an elegant solution to the problem. It has a unique advantage over the ambiguous biochemical strategies, that is one can be sure that the isolated gene is really involved and necessary in the biological process in question (see Csanadi et al,1994)