Gene Cloning and Characterization - Genetics, Genomics and Breeding of Crop Plants - page 179

6.1 Introduction
In sunflower, the large size of the genome (3,500 Mb; Baack et al. 2005) and
high content of repetitive DNA (e.g., Ty3- gypsy and Ty1- copia -like DNA;
Santini et al. 2002; Natali et al. 2006) pose a challenge for cloning genes of
agronomic importance. However, the development of new genomic resources
and databases from different genome projects, also for the Asteraceae family,
has made the sunflower genome more accessible to molecular analysis.
Using different techniques (Chapter 3), several types of molecular markers
have been detected that are closely linked to simple inherited traits
(Chapter 4) and placed on various genetic maps, constructed for sunflower
(Chapter 3). These markers and the positions of the genes on the genetic
maps represent the initial steps for isolating genes by map-based cloning
approaches. Even though markers can be very efficiently used in marker-
assisted breeding programs (Chapter 7), cloning of genes is necessary to
understand their function and regulation in different tissues and under
various environmental conditions. Map-based cloning or positional cloning
has not resulted in any isolation of genes in sunflower so far, but candidate
gene approach has been very successful and combination of map-based
cloning and candidate gene approaches might in the near future allow
successful isolation of additional genes in sunflower. In this chapter, we
summarize the attempts made to clone and characterize nuclear-encoded
genes in sunflower.
6.2 Map-based Cloning Approaches in Sunflower
Map-based cloning approach is used if the function of the gene of interest is
unknown and no corresponding gene has been isolated from another species.
Map-based cloning approach is reasonable only under these conditions
as it is extremely time-consuming. Map-based cloning requires the
identification of markers tightly linked to the gene of interest that can then
be used as probes against large genomic insert libraries to identify genomic
clones near the gene of interest or containing the gene of interest (Tanksley
et al. 1995). So, prerequisites for map-based cloning are: (1) availability of
large mapping populations, e.g., F 2 , RILs, F 1 BC 1 populations to develop high-
density genetic maps around the target locus, and (2) large-insert genomic
DNA libraries, e.g., BAC libraries. The latter have been and still are
representing a limiting factor in map-based cloning in sunflower as neither
the genome coverage nor the average insert sizes of the BAC libraries are
very satisfactory. This has and will hamper future map-based cloning
attempts in sunflower. Table 6-1 provides an overview on the available BAC
libraries. So far, all BAC libraries in sunflower except the library constructed
by Özdemir et al. (2004) have been made from maintainer lines. Apart from