Segregating populations derived from the cross [Lycopersicon esculentum x L. pennellii (LA 716)] have been used to determine the chromosomal locations of many isozyme and RFLP loci. However, good and consistent resolution is difficult to attain for the Prx-4 and Prx-7 alleles of these genotypes. The gene products of these loci are found in root tissue, and are resolved with the standard Paulik system (gel: tris 190 mM, citrate 50 mM, pH 7.8 with tris or citrate; electrode: boric acid, pH 7.9 with 4N NaOH; Rick et al., 1974). The only difference in zymotype between homozygous L. esculentum (+/+) and heterozygous (+/p) genotypes at Prx-4 band is frequently poorly resolved in this system. Resolution of the + and p alleles of the cathodal Prx-7 is also difficult with the Paulik system.
We have obtained significantly better resolution of the L. esculentum and L. pennellii (LA 716) alleles of Prx-4 and Prx-7 in the cathodal zone of histidine gels. Root extracts from young seedlings or two week old cuttings were electrophoreses in starch gels at constant voltage (13.3 V/cm) for 4h at 2_C using the Histidine/tris citrate gel/buffer system (gel: histidine_HCl 5 mM, pH 7.0 with 0.1N NaOH; electrode: tris 135 mM, citrate 43 mM, pH 7.0 with tris or citrate; Shields et al., 1983). The cathode portion of the gel was sliced and stained for 30-60 minutes at room temperature using a 3-amino-9-ethylcarbazole method (Vallejos, 1983).
Independent assortment analyses were conducted comparing segregating PRX bands observed in the cathodal zone of histidine gels with those observed in the Paulik system. In the Histidine system, the locus in the cathode portion of the gel nearest to the origin cosegregated with Prx-4, whereas the most cathodal locus cosegregated with Prx-7 (Fig. 1). The band corresponding to the Prx-4+ allele migrated approximately 1.4 cm; the Prx-4p allele migrated 4 mm behind (Fig. 1). The Prx-7+ allele migrated 3.0 cm from the origin, whereas, the Prx-7p allele was retarded 5 mm (Fig. 1). The resolution between + and p alleles at Prx-4 and Prx-7 in the Histidine system facilitated the genotypic classification of segregants at those two loci (Fig. 1).
Literature cited:
Rick, C.M. and S.D. Tanksley. 1981. Pl. Syst. Evol. 139:11-45
Rick, C.M., R.W. Zobel, and J.F. Fobes. 1974. Proc. Nat. Acad. Sci. 71:835-839.
Shields, C.R., T.J. Orton, and C.W. Stuber. 1983. In: Isozymes in plant genetics and breeding. pp 443-468. Elsevier Science Publishers, Amsterdam.
Vallejos, C.E. 1983. In: Isozymes in plant genetics and breeding. pp 469-516. Elsevier Science Publishers, Amsterdam.
Figure 1. Cathode portion of the histidine gel stained for peroxidase activity. 0> = origin; 4^c^> = cathodal Prx-4; 7> = Prx-7; e = Lycopersicon esculentum cv. Bonny Best; p = L. pennellii accession LA 716; and F = F1 of e x p. The other lanes are BC1 individuals from the cross e x (e x p).