germplasm at the tetraploid level that combines many good qualities of fused diploid selections. We have used this technique to greatly expand the tetraploid germplasm available for interploid crosses ( Grosser et al., 2010a , 2010b ). Used further in
Milica Ćalović, Chunxian Chen, Qibin Yu, Vladimir Orbović, Frederick G. Gmitter Jr and Jude W. Grosser
Jude W. Grosser, Hyun Joo An, Milica Calovic, Dong H. Lee, Chunxian Chen, Monica Vasconcellos and Frederick G. Gmitter Jr
interploid crosses for triploid production ( Grosser and Gmitter, 2009 ). In fact, citrus somatic hybridization has proven to be a key tool to generate allotetraploid breeding parents for use in interploid crosses to generate seedless triploids for mandarin
Irene E. Palmer, Thomas G. Ranney, Nathan P. Lynch and Richard E. Bir
, to evaluate self-compatibility and crossability between species, to determine interploid crossability among R. hirta cultivars, and to assess reproductive pathways in triploid R. hirta to better facilitate future Rudbeckia breeding projects
Charles J. Simon and John C. Sanford
A method is described for separating large and small pollen effectively from a heterogeneous mixture. This method potentially is applicable to separation of pollen grains of different ploidy levels, since “unreduced” 2n pollen is larger than normal pollen (n); it might then be used to increase the efficiency of a breeding program employing sexual polyploidization and to diminish crossing inefficiencies in interploid crosses.
Jason D. Lattier and Ryan N. Contreras
minimum of 15 resolved cells were investigated per taxa. Pollen cytology. From a previous study on cross-compatibility ( Lattier and Contreras, 2017 ), a single seedling from an interploid cross was discovered to have a larger genome size than either
J.W. Grosser, J. Jiang, F.A.A. Mourao-Fo, E.S. Louzada, K. Baergen, J.L. Chandler and F.G. Gmitter Jr.
Seedlessness is an important breeding objective of most citrus scion improvement programs, but production of quality seedless triploid citrus via interploid crosses has historically been limited by the low quality of available tetraploid parents. Production of tetraploid hybrid parents from elite diploid scion cultivars via protoplast fusion is now a practical strategy, and numerous hybrids can be produced on a timely basis from a wide range of parents. Such hybrids can be used as pollen parents in interploid crosses to generate improved seedless triploid fresh fruit cultivars. Herein we report the production of 15 such hybrids from 17 different parents, including sweet orange [Citrus sinensis (L.) Osbeck], mandarin/tangerine (C. reticulata Blanco), grapefruit (C. paradisi Macf.), pummelo [C. grandis (L.) Osbeck], tangor (C. reticulata × C. sinensis), and tangelo (C. reticulata × C. paradisi) germplasm. All hybrids were confirmed by cytological and RAPD analyses, and have been budded to selected rootstocks to expedite flowering.
N. Tusa, J.W. Grosser, F.G. Gmitter Jr. and E.S. Louzada
Allotetraploid somatic hybrid plants of `Hamlin' sweet orange (Citrus sinensis L. Osbeck) + `Femminello' lemon (C. limon L. Burm. f.), and Milam lemon (purported hybrid of C. jambhiri Lush) + `Femminello' lemon were regenerated via somatic embryogenesis following protoplast fusion. `Hamlin' and Milam protoplasts were isolated from undeveloped ovule-derived embryogenic callus cultures and fused using a polyethylene glycol method with seedling leaf-derived protoplasts of `Femminello' lemon. Somatic hybrids were identified on the basis of leaf morphology, root-tip cell chromosome number, and electrophoretic analyses of phosphoglucose isomerase, phosphoglucose mutase, and 6-phosphogluconate dehydrogenase leaf isozymes. The somatic hybrids will be used in interploid crosses with lemon in an effort to generate seedless triploid lemon types with improved tolerance to mal secco disease.
Jude W. Grosser
Citrus protoplast technology has advanced to where several practical applications in variety improvement and plant pathology are routine. We will report on progress in the following areas: somaclonal variation—`Valencia` and `Hamlin' sweet orange protoclones have been selected for improved juice color, higher soluble solids, seedlessness, and altered maturity dates; somatic hybridization for scion improvement—allotetraploid breeding parents have been created from numerous combinations of elite parental material, and are now being used as pollen parents in interploid crosses to produce seedless triploid varieties; somatic hybridization for rootstock improvement—numerous somatic hybrids combining complementary rootstock germplasm are under commercial evaluation and several look promising for wide adaptation, improved disease resistance, and tree size control; transformation—an alternative protoplast-based transformation that utilizes EGFP for selection has been developed; virus resistance assays—a protoplast-based assay is being used to screen varieties and candidate sequences for resistance to citrus tristeza virus at the cell level, saving time and greenhouse space.
Joseph J. Rothleutner, Mara W. Friddle and Ryan N. Contreras
The genus Cotoneaster (Rosaceae, Maloideae) is highly diverse, containing ≈400 species. Like other maloids, there is a high frequency of naturally occurring polyploids within the genus, with most species being tetraploid or triploid. Apomixis is also prevalent and is associated with polyploidy. The objective of this study was to estimate genome sizes and infer ploidy levels for species that had not previously been investigated as well as compare estimates using two fluorochromes and determine base pair (bp) composition. Chromosome counts of seven species confirmed ploidy levels estimated from flow cytometric analysis of nuclei stained with 4′,6-diamidino-2-phenylindole (DAPI). Monoploid (1Cx) genome sizes ranged from 0.71 to 0.96 pg. Differences in monoploid genome size were not related to current taxonomic treatment, indicating that while chromosome sizes may vary among species, there are no clear differences related to subgeneric groups. A comparison of DAPI and propidium iodide (PI) showed a difference in DNA staining in Cotoneaster comparable to other rosaceous species. Base pair composition (AT%) in Cotoneaster ranged from 58.4% to 60.8%, which led to overestimation of genome size estimates in many cases—assuming the estimates of the DNA intercalator are accurate. Our findings will inform breeders with regard to the reproductive behavior of potential parents and may be used to confirm hybrids from interploid crosses.
Jude Grosser, Milicia Calovic, Patricia Serrano, Fred Gmitter Jr. and J. L. Chandler
The international fresh citrus market now demands high-quality, seedless fruit that must also be easy to peel for consumer convenience, especially when considering new mandarin varieties. High quality varieties that historically perform well in Florida are generally seedy. Florida is therefore losing market-share to `Clementine' and other seedless varieties produced in Mediterranean climates, including Spain, Morocco, and California. In our ongoing program, somatic hybridization and cybridization via protoplast fusion are now playing a key role in strategies to develop competitive seedless mandarin hybrids adapted to Florida. Somatic hybridization is being used to combine elite diploid parents to produce high quality allotetraploid breeding parents that can be used in interploid crosses to generate seedless triploids. Several thousand triploid mandarin hybrids have been produced under the direction of F.G. Gmitter, Jr. Some of our allotetraploid somatic hybrids are producing fruit with direct cultivar potential, i.e., 'Valencia' sweet orange + `Murcott' tangor. New somatic hybrids produced in our program will be discussed, including `Page' tangor + `Dancy' mandarin, `Page' tangor + `Kinnow' mandarin, and `Hamlin' sweet orange + LB8-9 tangelo. Somatic cybridization is being used to transfer CMS (cytoplasmic male sterility) from the seedless `Satsuma' mandarin to other seedy varieties via mtDNA transfer, in efforts to make them seedless. New somatic cybrids produced in our program that contain the `Satsuma' CMS include `Murcott' tangor and `Kinnow' mandarin. Details of these results and other progress will be discussed.