Cucurbit Genetics Cooperative Report 23: 49-50 (article14) 2000
A. Sanchez-Donaire; J.M. Guerra-Sanz
C.I.F.H. Almeria P.O. Box 91 El Ejido (Almeria, -Spain)
C.L. Encina
Est. Exp. “La Mayora”. CSIC, Algarrobo-Costa (Malaga.-Spain)
Introduction: Diploid watermelon (Citrullus lanatus (Thunb.) Matsum. & Nakai) genotypes were screened in order to determine the best explant material, as well as genotypes, for in vitro regeneration. This information will help other researchers who are conducting genetic transformation studies in watermelon, or any other in vitro manipulation, such as in vitro selection.
Two in vitro explant regeneration systems have been reported: one from juvenile cotyledons (1), and the other from immature cotyledons (2). Both systems use similar culture media, and regeneration is by adventitious shooting. For juvenile cotyledons, explants are taken five days after germination, however for immature cotyledons, explants are taken from fruits 28 days after pollination.
Materials and Methods: Sixteen genotypes of watermelon were used in this study, including commercial varieties, breeding lines and landraces. Their names and origin of each genotype are listed in Table 2.
At least 30 explants of each genotype and assay (juvenile or immature cotyledons) were used to evaluate in vitro regeneration. Regeneration was considered to have occurred if at least one shoot from an explant had developed by six weeks post-induction.
In vitro culture were observed after four and six weeks, and the following data were collected: explant number per genotype; number of callusing explants; number of explants with structures potentially regenerative; and number of shooting explants.
Analysis of Variance (ANOVA) was performed with each explant considered a replication (Table 1). The results are reported as the percentage of regenerated explants per genotype at six weeks post-organogenesis (Table 2).
Results: The ANOVA (Table 1) shows that there are differences among genotypes, as well as between the source of the explants. Comparisons among genotypes for the percentage of regenerable explants are found in Table 2.
The results found in Table 2, suggest that the best genotype for regeneration LPKKA when considering both sources of explants. LPKKA had 71.4% regenerated explants from juvenile cotyledons and 100% from immature cotyledons. PLKKA is followed by the landraces LG7 and LG8 when considering only the juvenile explants. LG8 had lower regeneration percentage with immature cotyledon. LPKKA is a breeding line, and therefore, could be used directly to introduce a trait into elite germplasm. The results from the landraces indicate that there is a great amount of available variability for in vitro selection that could be used in a crossing program.
In general, the best explant for regeneration was the immature cotyledon (Table 2). However, from a practical point of view, it is important to note that the immature cotyledon explant system, requires growing the plant past fruit setting, whereas juvenile cotyledon explants can be obtained after only two weeks growth.
Table 1. Analysis of Variance – Type III sums of Squares
| Source | Sums of squares | Df | Mean squares | F-ratio | P-value |
| Main effects | |||||
|
2817.99 | 15 | 187.866 | 2.25 | 0.0135 |
|
72915.2 | 1 | 72915.2 | 872.30 | 0.0000 |
| Interaction (G X E) | 1524.06 | 15 | 101.604 | 1.22 | 0.2849 |
| Residual | 5182.53 | 62 | 83.5892 | ||
| Total (corrected) | 95630.2 | 93 | |||
Table 2. Percentage of regenerated explants
| Genotype | Origin | Juvenile cotyledon | Immature cotyledon |
| DULCE | Novartis, S.A. | 15.3 Bx | 94.4 bed |
| MARAVILLA | Ramiro Arnedo | ||
| PANONIA | Seminis | 33.3 abc | 100 d |
| PATA NEGRA | Seminis | 16.1 ab | 100 d |
| LPKKA | Fito | 71.4 c | 100 d |
| LP3B | Las Gabias | 18.45 ab | 97.2 cd |
| LG4z | Las Gabias | 8.33 ab | 84.72 ab |
| LG6 | Las Gabias | 8 a | 91.66 bcd |
| LG7 | Las Gabias | 45.11 bc | 100 d |
| LG8 | BGHy | 33.3 bc | 83.3 abcd |
| L12899 | BGH | 24.64 bc | 100 d |
| L12900 | BGH | 18.2 ab | 88.8 abcd |
| L12903 | BGH | 20.31 abc | 100 cd |
| L12904 | BGH | 13.69 ab | 100 cd |
| L12907 | BGH | 23.47 bc | 100 d |
| L12911 | BGH | 30.35 bc | 77.7 a |
| L12917 | BGH | 15.4 ab | 100 d |
z LG = Watermelon landraces from Las Gabias (Granada).
y BGH = Watermelon landraces from Horticulture germplasm, Spanish Bank (F. Nuez; Univ. Politecnica de Valencia. – Valencia Spain).
x Different letters indicate statistically significant differences between genotypes, P = 0.05% after LSD test.
Acknowledgements: We appreciate the help of all the seed companies cited in Table 2 for their generous contributions of the genotypes used in this study. We thank Emila Romero for her help in the laboratory and field experiments. This project was partially funded by Fundacion para la Investigciou de la Provincia de Almeria (FIAPA).
Literature Cited
- Compton, M.E. and D.J. Gray, D.J. 1993. shoot organogenesis and plant regeneration from cotyledons of diploid, triploid and tetraploid watermelon. J. Amer. Soc. Hort. Sci. 118:151-157.
- Zhang, X.P., B.B. Rhodes and J.W. Adelberg, 1994. Shoot regeneration from immature cotyledon of watermelon. CGC Rep. 17: 36-39.