No abstract

We have developed molecular markers tightly linked to Sex1, the gene that determines plant sex in papaya ( Carica papaya L.). Three RAPD products have been cloned and a portion of their DNA sequenced. Based on these sequences SCAR primers were synthesized. SCAR T12 and SCAR W11 produce products in hermaphrodite and male plants and only rarely in females. SCAR T1 produces a product in all papayas regardless of plant sex. SCAR T12 and SCAR W11 showed no recombination in a population of 182 F2 plants from a 'SunUp' by 'Kapoho' cross. Based on these results a PCR-based technique for rapidly and accurately determining the sex of papaya plants was developed using either W11 or T12 to detect the hermaphrodite or male allele and T1, which amplifies a product regardless of sex type, as a positive control. The sexing technique, using SCAR T12 and SCAR T1 as a positive control, was used to correctly predict hermaphrodite papaya plants in a population of seedlings with an overall accuracy of 99.2%.

A high-density genetic map of papaya (Carica papaya L.) was constructed using 54 F 2 plants derived from cultivars Kapoho and SunUp with 1501 markers, including 1498 amplified fragment length polymorphism (AFLP) markers, the papaya ringspot virus coat protein marker, morphological sex type, and fruit flesh color. These markers were mapped into 12 linkage groups at a LOD score of 5.0 and recombination frequency of 0.25. The 12 major linkage groups covered a total length of 3294.2 cM, with an average distance of 2.2 cM between adjacent markers. This map revealed severe suppression of recombination around the sex determination locus with a total of 225 markers cosegregating with sex types. The cytosine bases were highly methylated in this region on the basis of the distribution of methylation-sensitive and -insensitive markers. This high-density genetic map is essential for cloning of specific genes of interest such as the sex determination gene and for the integration of genetic and physical maps of papaya.

Genetic relationships among Carica papaya cultivars, breeding lines, unimproved germplasm, and related species were established using amplified fragment length polymorphism (AFLP) markers. Seventy-one papaya accessions and related species were analyzed with nine EcoRI-MseI primer combinations. A total of 186 informative AFLP markers was generated and analyzed. Cluster analysis suggested limited genetic variation in papaya, with an average genetic similarity among 63 papaya accessions of 0.880. Genetic diversity among cultivars derived from the same or similar gene pools was smaller, such as Hawaiian Solo hermaphrodite cultivars and Australian dioecious cultivars with genetic similarity at 0.921 and 0.912, respectively. The results indicated that self-pollinated hermaphrodite cultivars were as variable as open-pollinated dioecious cultivars. Genetic diversity between C. papaya and six other Carica species was also evaluated. Carica papaya shared the least genetic similarity with these species, with an average genetic similarity of 0.432; the average genetic similarity among the six other species was 0.729. The results from AFLP markers provided detailed estimates of the genetic variation within and among papaya cultivars, and supported the notion that C. papaya diverged from the rest of Carica species early in the evolution of this genus.

Since 1992, Papaya ringspot virus (PRSV) destroyed nearly all of the papaya hectarage in the Puna district of Hawaii, where 95% of Hawaii's papayas are grown. Two field trials to evaluate transgenic resistance (TR) were established in Puna in October 1995. One trial included the following: SunUp, a newly named homozygous transformant of Sunset; Rainbow, a hybrid of SunUp, the nontransgenic Kapoho cultivar widely grown in Puna, and 63-1, another segregating transgenic line of Sunset. The second trial was a 0.4-ha block of Rainbow, simulating a near-commercial planting. Both trials were installed within a matrix of Sunrise, a PRSV-susceptible sibling line of Sunset. The matrix served to contain and trace pollen flow from TR plants, and as a secondary inoculum source. Virus infection was first observed 3.5 months after planting. At a year, 100% of the non-TR control and 91% of the matrix plants were infected, while PRSV infection was not observed on any of the TR plants. Fruit production data of SunUp and Rainbow show that yields were at least three times higher than the industry average, while maintaining percent soluble solids above the minimum of 11% required for commercial fruit. These data suggest that transgenic SunUp and Rainbow, homozygous and hemizygous for the coat protein transgene, respectively, offer a good solution to the PRSV problem in Hawaii.