SUNFLOWER GERMPLASM DIVERSIFICATION AND CHARACTERIZATION UTILIZING WILD SUNFLOWER SPECIES, CYTOGENETICS, AND APPLIED GENOMICS
Location: Sunflower Research
Title: Genetic analysis and molecular mapping of an Rf gene from Helianthus angustifolius for a new cytoplasmic male-sterile line
Submitted to: Meeting Abstract
Publication Type: Abstract Only
Publication Acceptance Date: December 21, 2011
Publication Date: February 26, 2012
Citation: Liu, Z., Wang, D., Jan, C. 2012. Genetic analysis and molecular mapping of an Rf gene from Helianthus angustifolius for a new cytoplasmic male-sterile line. In: 18th International Sunflower Conference Program and Abstracts, Mar del Plata & Barcarce, Argentina, February 27-March 1, 2012. P.234.
The combination of cytoplasmic male-sterile (CMS) and the corresponding fertility restoration genes (Rf) is a critical tool in large-scale hybrid seed production of sunflower. A new CMS line 514A, derived from H. tuberosus / 7718B, was obtained from a scientific exchange with the Liaoning Academy of Agricultural Sciences, Liaoning, China. In order to identify its Rf genes, 33 maintainer and restorer lines from five countries for CMS PET-1 cytoplasm and 20 tester lines from USDA-ARS-NCSL were used, but none of them restored the fertility for CMS 514A. The aims of this study were: (1) to identify the Rf gene using different materials; (2) to analyze the genetics of the Rf gene; and (3) to locate the Rf gene on a molecular linkage map. Crossing and backcrossing method were used for detecting the Rf genes and reducing the chromosome number to 2n=34. Genomic in situ hybridization (GISH) was used to verify the alien chromosome or segments. Two F2 populations were used for mapping the gene. The F3 progeny test was used to phenotype the F2 individuals. Bulk segregation analysis was used to screen the 370 SSR markers spanning the 17 sunflower linkage groups. An additional SSR and EST-SSR markers from the candidate linkage group (LG) were employed for mapping the gene. The deviation analyses of the fertility trait and marker loci were compared with the expected Mendelian ratios in the F2 generation using the Chi-square test. The MAPMAKER/Exp version 3.0b software was used for linkage analysis, with the Kosambi mapping function. Five amphiploids and one tetraploid were crossed to CMS 514A, with the F1 progenies pollinated by one amphiploid (Amp H. angustifolius / P 21, 2n=68) that produced all male-fertile plants with 2n=51. After several backcrosses with HA 89, male-fertile plants with 2n=35 were obtained. Genetic study of two male-fertile plants (2n=35) suggested that the Rf gene was located on the alien chromosome. The fertile plants with 2n=34 were obtained after backcrossing. The alien chromosome or segments were detected by GISH. Molecular mapping located this Rf gene on LG 3 of the public sunflower SSR map. Eight markers were linked to this gene, covering a genetic distance of 19.8 and 11.7 cM in the two mapping populations, respectively, with the closest marker ORS13 at a distance of 2.8 cM. The Rf gene was identified in the amphiploid of H. angustifolius / P 21. The Rf gene is expected to be on one alien chromosome in the fertile progenies with 2n=35. The Rf gene has been transferred into the progenies with 2n=34 using the backcross method. The gene was mapped to LG 3 of the sunflower map. The closely linked marker can be used for marker-assisted selection for this Rf gene. The study provides a new CMS/Rf gene system. Further analysis of the alien segment in the fertile F2 individuals will be conducted using the GISH technique. Meiotic analysis will also be used for chromosome pairing study in the future. A single CMS PET-1 originated from wild Helianthus petiolaris subsp. petiolaris Nutt. has been widely used for commercial sunflower hybrid seed production since it was discovered in1969. The maintainer and restorer lines for CMS PET-1 failed to restore the fertility of CMS 514A, suggesting this CMS is different than CMS PET-1. The Rf gene was finally detected in an amphiploid of H. angustifolius / P 21, and was located on LG 3. The results provide an alternative CMS/Rf gene system, which will expand the diversity of sunflower germplasm for breeding, and help to understand the mechanism of the interaction of cytoplasm and nuclear genes.