The fa2 gene and molecular markers mapping in the gp segment of the Pisum linkage group V

Review studies on the world Pisum genetic resources have shown that stem fasciation is controlled by three loci, i.e., fa1 (LGIV; Wt 10006 - type line of the Polish Gene Bank), fa2 (LGV, the line Wt 12185), and fas (LGIII, the line Shtambovii). Outstanding advantages of this character (e.g., pods gathered in upper part of a stem) resulted in breeding some cultivars. Preliminary investigations suggested linkages of the newly described fa2 gene within the gp–U segment. Based on the further linkage test crosses, it was stated that the fa2 is localized between the gp and Pis_Gen_9_3_1 markers (in the LGV). Additionally, four molecular markers (AD175, AB146, AC58, and AD280) and the morphological marker lk were also localized in this segment. Moreover, rms5, lum3, and cri were found to map on the other side of gp with tight linkage observed between lum3 and cri.


Introduction
Stem fasciation in peas appears to be a very interesting character from a theoretical as well as a practical point of view. This character not only changes the stem architecture but also the physiology of flowering and maturing (Fig. 1). Its advantage is that pods are gathered in upper part of a stem; but in consequence, pea plants lodge and are susceptible to drought during shortened flowering and pod formation periods. Outstanding advantages resulted in breeding some cultivars, for example, cvs. Buława (POL), Ornamenta, Rosacrone, Golf (DEU) and Novella (USA). Pea fasciation was described for the first time in 1597 (Święcicki 2001(Święcicki , after Derbshire 1911, and since then different names have been used for its designation, such as the Pisum umbellatum, mummy pea, or crown pea. Furthermore, a taxon was separated in Pisum taxonomy, i.e., P. sativum, subsp. sativum, convar. vulgare var. coronatum (Lehmann and Blixt 1984).
Fasciation was one of the seven monohybrid characters studied by Mendel (Święcicki et al. 2000). Induced mutations resulted in a number of independent mutation cases with a similar phenotype in different genotypic backgrounds which are still maintained in world Pisum collections (e.g., USDA Pullman Pisum Genetic Stock Collection, John Innes Centre Pisum Collection, Wiatrowo Pisum Collection). Information on the character anatomy, morphology, and expression are available in several references such as Gottschalk and Wolf (1983), Marx and Hagedorn (1962) and Sinjushin and Gostimsky (2006), but different opinions are available on its mode of inheritance. It has been shown that this character is controlled by one to four independent genes or multiple alleles of a single locus (Marx and Hagedorn 1962;Blixt 1972;Lamprecht 1974;Święcicki 2001). The most popular was the acceptance of two independent fasciata genesfa in LG IV and fas in LG III (Lamprecht 1974;Blixt 1977). Additionally, a similar mutation type, dichotomous branching, was selected and reported as a character governed by two polymeric genes bif1 and bif2 (Gottschalk and Wolf 1983); this alteration was associated with a fasciation of only a few upper nodes that results in a forked stem (Fig. 2). Results of subsequent complementation tests (locus identity test crosses) that explain the genetic basis of fasciata phenotype in pea lines from the Blixt's, Gottschalk's, Marx's, Święcicki's and Sinjushin's collections were as follows (Święcicki 2001;: no typeline registered by Blixt (1977)

exists in the main
Pisum collections for the fas gene from LGIII;  and Sinjushin (2011) suggested that the two lines, JI2771 and mutant Shtambovii, have the fas gene, fasciation in most of the tested lines is controlled by the fa gene from LGIV, dichotomous branching appeared to be controlled by the allele in the fa locus (symbol fa bif was suggested), an exception is the fasciation in the accession Wt 12185 as controlled by a gene different from the fa locus; for fa in LGIV, the symbol fa1 (and fa1 bif ) was suggested and fa2 for the new gene in the type line Wt 12185.
For usage of a stem fasciation in breeding, the results of Gottschalk (1979) and Święcicki (2001) are important, indicating that lines Gott37B (fa1 bif ) and Wt12185 (fa2) are characterised by a full penetrance of mentioned genes and an increased seed production.
Preliminary results have shown that the new fasciata gene, fa2, is linked with gp in linkage group V (LGV) (Cr0 = 17.6 ± 7.6) (Święcicki and Gawłowska 2004). The aim of this study was to map fa2 using more markers from the gp region, including molecular markers.
SSR primers were designed by the Pea Microsatellite Consortium, Agrogene, France, and used in mapping by Loridon et al. (2005). The results of mono-and dihybrid segregations were calculated using a computer program based on the product-ratio method for linkage estimation (Święcicki et al. 1998). For graphic presentation of the loci order, the MapChart program was used (Voorrips 2002).

Results and discussion
Mono-and dihybrid segregations in the F 2 generation of seven populations were analysed by the fa2 gene and eight morphological and five molecular markers (Table 1). Preliminary results suggest that most of the selected markers originate from the Gp-Fa2-U region (Święcicki and Gawłowska 2004). Markers lum3, cri, and cp-1 localized on the opposite side of gp (Blixt 1977;Święcicki 1988;Weeden et al. 1998) additionally should confirm a selection of appropriate chromosome region for the fa2 localization. And it appeared ( Correct, monohybrid segregation for the fa2 and selected markers (Table 1) allowed us to analyse a dihybrid segregation to look for linkages and the fa2 locus (Table 2). In K. 3548 and K. 3048 populations  Table 2) additionally confirm the fa2 localization and the presented loci order (see also consensus Pisum map, Weeden et al. 1998). A valuable supplement for this region is the linkage Fa2-Lk = 13.8, revealed in the population K. 3364. Together with the earlier result for Gp-Lk = 12.3 (Święcicki 1989), it is emphasized that the Lk gene is also localized in the investigated Gp-U region. Supplemental linkage data in the above-mentioned region supply an analysis of K. 3319 and K. 3338 populations covering the Gp locus and five molecular markers: Pis_Gen_9_3, AD175, AB146, AC58, and AD280. Obtained results suggest the loci order given in Fig. 3.
Conducted analyses localized the new fa2 gene in the Gp-U segment of the LGV between Gp and Pis_Gen_9_3 markers. Four additional molecular markers (AD175, AB146, AC58, AD28) and morphological lk were also localized in this segment. Moreover, the locus rms5 and a strong linkage between lum3 and cri were found from the other side of the Gp locus.