Molecular characterization of chemical mutagenesis induced diversity in elite maize germplasm

摘要

Three classical breeding Iowa Super Stiff Stalk (SSS) inbred lines B37, B73 and B84, one Lancaster inbred Oh43 and mutant lines obtained by chemical mutagenesis followed by mutation breeding as follows: two of B37 and four of Oh43 were selected for molecular characterization. The mutant inbred lines were chosen because in addition to the improved GCA and SCA for grain yield, proven by their predominance in the Bulgarian breeding programs, they showed shifts in the flowering time as compared to the initial inbreds. Molecular markers (micro satellites and other PCR-based DNA markers) were used for characterization of maize genotypes and determination of the induced by chemical mutagenesis genetic variability in maize germplasm. The tested nine SSR markers (umc 1001, umclO14, umcl057, umcll81, umcl0lS, umc 1029. umcl003, umc 1033 and umcl035) can discriminate between the initial classical breeding inbred lines and the originating mutant inbreds. Allelic diversity was also studied by PCR amplification with specifically de-signed primers in the coding regions and flanking sequence of two genes: dwarf8 (d&: chromosome 1, 198.5 cM), and indeterminate l (id1; chromosome 1. 175.0 cM). These are considered candidate genes for variation in plant height and/or flowering time, based on mutant phenotypes and chromosomal locations near major QTLs. Single nucleotide polymorphisms and indels were detected in the region flanking the SH2 domain of dwarf8 gene in some of the mutant inbreds as a result of SSCP and sequencing analyses. However, these polymorphisms could not be associated with the observed variations in flowering time. PCR analysis of the promoter region dwarf8 showed a variant fragment of about 1 kb in the inbred line Oh43 that was not present in any other initial and mutant in-bred lines included in the study. PCR amplification of the 5' end of the Id1 coding sequence revealed polymorphic bands in the mutant lines XM535, XM521, XM250-l, XM98-8 and XM85-105, as well as in the classical breeding line B73. The data, presented here demonstrate the usefulness of chemical mutagenesis for generation of genetic diversity within the elite maize germplasm. Some of this variation may affect the major genes in the QTLs. Our initial data revealed mutagenesis induced polymorphisms in the coding sequences of two important for the determination of flowering time transcription factors. Further molecular analyses of the proposed model systems may complement the trait association efforts and will help to directly identify the major genes in the QTLs.