History

• Early agriculturalists selected natural Citrus Breeding hybrids and mutants for seed propagation

• 1800’s- grafting and cuttings became popular to propagate best varieties Kevin M. Crosby • 1900’s- artificial cross-pollination practiced

Breeding Goals Rootstock

• Rootstock- tree size, stress tolerance • Dwarfing- Poncirus, some mandarins

• Scion- fruit color, size, shape, flavor, yield • Compatability- citrus better than Poncirus

• Disease resistance- CTV, Phytophthora, • Seedling vigor and scion yield CVC, Alternaria, Scab, Greening, etc. • Fruit quality- size, shape, flavor, juice

Rootstock Hybrids Carrizo Citrange

• ‘Carrizo,’ ‘Troyer’ Citrange- navel orange x Poncirus, very popular in FL, CA

• ‘Swingle’ Citrumelo- grapefruit x Poncirus, very popular in Florida, salt intolerant

• ‘Sunki’ x ‘Swingle’ tf- semi-dwarfing, salt- tolerant, possible replacement for SO in TX

1 Swingle Citrumelo Rootstock Fruit

Scion Scion Hybrids

• Vigor, yield potential, cold tolerance • ‘Orlando,’ ‘Minneola’ tangelos- ‘Duncan’ grapefruit x ‘Dancy’ tangerine • Fruit type- mandarin grapefruit, orange • ‘Page’ mandarin- ‘Minneola’ x ‘Clementine’ • Fruit quality- flavor, size, seediness, appearance, shelf-life • ‘Oro Blanco’ grapefruit triploid- tetraploid pummelo x grapefruit

Disease Resistance Breeding Techniques

• Viruses- CTV, Psorosis, Exocortis • Cross-pollination- combine genes from different parents in hybrid progeny • Bacteria- Citrus Variegated Chlorosis, Greening, Canker • Self-pollination- fix genes of interest in one line to stabilize phenotype (inbreeding)

• Fungi- Alternaria, Scab, Melanose, • Mutation- natural or induced genotypic Phytophthora modification

2 Cross Pollination Self-pollination

• Combine desirable traits from different • Natural mechanism for species to maintain genotypes/species and exploit heterosis genetic uniformity- pummelo, mandarins

• Swingle- one of earliest to make extensive • Serious inbreeding depression in citrus crosses for rootstock improvement overcome by apomixis- nucellar embryony

• Many hybrid rootstocks between Poncirus • Important for gene inheritance and function and Citrus- citrange, citrumelo, citrandarin studies

Seed Structure Mutation

• Natural mutations- ‘sports’ of buds or limbs: ‘Ruby Red,’ most orange varieties

• Gamma rays- chromosome breaks cause genotypic changes: ‘Star Ruby,’ ‘Rio Red’

• Chemical and t-DNA- interrupt single genes

Population Development Pedigree Method

• Pedigree- all progeny from specific cross • Most citrus varieties developed by this carefully evaluated, limited genetic base method- few crosses

• Recurrent selection- diverse populations • Relatively few parents (monoembryonic) as improved by selection and intercrossed females; various males

• Mass selection- large population evaluated for a few outstanding individuals • Each progeny evaluated from each family

3 Recurrent Selection Mass Selection

• Each population developed for important • Characterized or heterogeneous populations traits- good genetic diversity from relatively few crosses screened

• Crosses between individuals from improved • Focus mainly on quantitative traits or genes populations evaluated for superior traits with incomplete penetrance

• Most productive over long period • Labor intensive but rapid improvement

Biotechnology Protoplast Fusion

• Protoplast fusion of different genotypes • Isolate cell protoplasts from callus or leaf tissue and fuse in vitro to form hybrids • Gene mapping with molecular markers- gene cloning. • Mostly polyploid plants regenerated from tissue culture- genetic hybrids • Genetic transformation with novel genes to modify DNA- Agrobacterium, biolistics • Overcome barriers to sexual reproduction

Gene Cloning Map-based Cloning

CGTTGA- part of FR gene • Mapping genes in DNA with molecular 120 kb BAC markers- RAPD, RFLP, AFLP, etc. RFLP but no RAPD

• Chromosome walking- locate DNA markers SCAR RFLP RAPD AFLP adjacent to gene of interest, clone gene inside bacterial plasmid 1 CM .4 .6 FINGERBLIGHT • cDNA cloning- isolate genes from mRNA RESISTANCE GENE

4 Genetic Transformation Texas Priorities

• Insertion of cloned gene sequence into • Salt and drought tolerance DNA (genome) of desirable plant • CTV and Phytophthora resistance • Modify single trait while maintaining good attributes of parent- SO with Ctv gene • Cold and heat tolerance

• Insertion point in genome not targeted • Fresh market fruit- size, sugars, low acid

Past Achievements in Texas Past Achievements in California

• ‘Ruby Red Grapefruit’- bud sport of • Hybrid mandarins- ‘Kinnow,’ ‘Pixie’ Thompson in LRGV, changed market • Triploid seedless grapefruit- ‘Oroblanco’ • ‘Star Ruby’- irradiated seedling of ‘Hudson,’ darkest red grapefruit • Hybrid red pummelo- ‘Chandler’ • ‘Rio Red’- irradiated budwood of ‘Ruby Red,’ most popular red grapefruit today • Rootstocks- citranges, citrumelos

Past Achievements in Florida Current Variety Development

• Tangelos- ‘Orlando,’ ‘Minneola,’ ‘Page,’ • New triploid, seedless mandarins- CA, FL

• Grapefruit- ‘Marsh seedless,’ ‘Duncan’ • New salt tolerant, dwarf rootstocks- CA,TX

• Tangors- ‘Murcott,’ ‘Temple,’ ‘Fallglo,’ • New low acid grapefruits- CA ‘Ambersweet’ • New fusion product rootstocks- FL

5 Current Molecular Research Future Goals

• CTV resistance gene cloning- CA,TX,FL • CTV resistance gene in susceptible scions and rootstocks • Low acid gene mapping- CA • Phytopthora resistant, salt tolerant, high • Fruit development gene mapping- CA,FL yielding rootstocks

• Chromosome Isolation and Fusion-TX • Fruit- sweeter, seedless, longer shelf life

Breeding Strategy for Texas

• Increase effort in transgenics development

• Increase emphasis on fruit quality and earliness for fresh market expansion

• Increase research into genetic cold tolerance

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