CYTOLOGICAL ASSESSMENT OF DOUBLED HAPLOIDS IN SUMMER SQUASH (CUCURBITA PEPO L.)

147 new forms were obtained only from nine responsive unpollinated ovules of summer squash in 2015-2017. The cytological analysis is used to estimate the level of ploidy in regenerated plants R0. The small size of mitotic chromosomes and large number in Cucurbita genus make it difficult to count them even though they are well separated. As a result of analysis the optimal method of chromosome staining has been chosen with the use of modified propiono- lacmoid cytological technique. The rootlet tips and apical meristems were used to make smear preparations. As it was shown the summer squash was a difficult species as a cytological object because of low mitosis frequency and low number of metaphase plates with well scattered chromosomes. The photos of chromosomes in squash C. pepo subsp. brevicaulis var. giraumons Duch; distant hybrid (breeding accession N37) between variety ‘Cornishon’ and winter squash C. pepo subsp. longicaulis Greb. var. pepo and doubled haploid plants produced from them were made. In spite of the small size 2 μm the chromosomes observed were clearly seen. Nearly all regenerated plants that had been analyzed passed well the adaptation in vivo and occurred to be a doubled haploids (2n = 2x=40). The seed progeny was then obtained through self-pollination. About 20% of plants R0 analyzed were mixploids and supposedly only 7% were haploids.

1. Vavilov N.I. Intergeneric hybrids of melons, watermelons and squashes. Proceedings on Applied Botany, Genetics and Breeding. V.14 (1). P.1-35.

2. Domblides E.A., Shmykova N.A., Zayachkovskaya T.V., Khimich G.A., Korotseva I.B., Kan L.Y. and Domblides A.S. Doubled haploid plant production from in vitro culture of unpollinated ovules of summer squash (Сucurbita pepo L.). Proceedings of the of the VII International Scientific and Practical Conference «Biotechnology as an Instrument for Plant Biodiversity Conservation (physiological, biochemical, embryological, genetic and legal aspects)», devoted to the 30th anniversary of Biotechnology Department in Nikita Botanical Gardens Yalta, Crimea, Russia September, 25 October, 01, 2016. Simferopol: PP «ARIAL», 2016. 352 p.

3. Domblides E.A., Shmykova N.A., Shumilina D.V., Zayachkovskaya T.V., Mineykina A.I., Kozar E.V., Akhramenko V.A., Shevchenko L.L., Kan L.Yu., Bondareva L.L., Domblides A.S. Technology of doubled haploid plant production through isolated microspore cultivation in Brassicaceae family. Federal State Budgetary Scientific Research Institution «All-Russian Scientific Research Institute of vegetable breeding and seed production», Moscow, 2016 (In Russ.).

4. Domblides E.A., Shmykova N.A., Khimich G.A., Korotseva I.B., Domblides A.S. DEVELOPMENT OF ABNORMAL FLOWERS IN GENERATION OF DOUBLED HAPLOIDS OF SUMMER SQUASH (CUCURBITA PEPO L.). Vegetable crops of Russia. 2018;(5):13- 17. (In Russ.) DOI:10.18619/2072-9146-2018-5-13-17

5. Kan L.Yu. Cytological studies in breeding work based on interspecific hybridization in the Allium genus. “Chromosomes and evolution”. Symposium in memory of G.A. Levitsky. St. Petersburg, 2008. P.53-55.

6. Pausheva Z.P. Laboratory manual on plant cytology / Z.P. Pausheva. Ed. 4th, pererabot. and add. Moscow: Agropromizdat, 1988. 271 p.

7. Puhalsky V.A., Soloviev A.A., Badaeva E.D., Yurtsev V.N. Workshop on cytology and plant cytogenetics. M.: Publishing house "Kolos-s". 2007.

8. Solovyova L.V. Laboratory manual on the fruit plant cytology. M.: Publishing House of Moscow State University, 1982. 54 p.

9. Timin N.I., Pyshnaya O.N., Agafonov A.F., Mamedov M.I., Titova I.V., Kan L.Yu., Logunova V.V., Romanov V.S., Shmykova N.A., Timina L.T., Gurkina L.K., Suprunova T.P., Krivosheev S.M., Engalycheva I.A. Interspecial hybridization in onion (Allium L.), carrot (Daucus L.), and pepper (Capsicum L.). Moscow, 2013.

10. Shmykova N.A., Suprunova T.P. (2009) An induction gynogenesis in culture in vitro not-pollinated seedbuds of Cucumis sativus L. Gavrish, 2009, 4: 40-44.

11. Shmykova N.A., Shumilina D.V., Kushnereva V.P., Khimich G.A. Gynogenesis induction in culture of unpollinated ovules of pumpkin. Vegetable crops of Russia. 2011. №1 (10). P.28-31. (In Russ.)

12. Shmykova N.A., Khimich G.A., Korotseva I.B., Domblides E.A. Prospective of development of doubled haploid plants of Cucurbitaceae family. Vegetable crops of Russia. 2015;(3-4):28-31. (In Russ.) https://doi.org/10.18619/2072-9146-2015-3-4-28-31

13. Chen, J., Staub, J. E., & Jiang, J. (1998). Genetic Resources and Crop Evolution, 45(4), 301–305. doi:10.1023/a:1008603608879

14. Domblides E.A., Shmykova N.A., Shumilina D.V., Zayachkovskaya T.V., Vjurtts T.S., Kozar E.V., Kan L.Yu., Romanov V.S., Domblides A.S., Pivovarov V.F., Soldatenko A.V. Biotechnological approaches for breeding programs in vegetable crops. Agrosym 2017. Book of proceedings. 2017. P.452-460.

15. Jeffrey C. (1980). A review of the Cucurbitaceae. Botanical Journal of the Linnean Society, 81(3), 233–247. doi:10.1111/j.1095-8339.1980.tb01676.

16. Kurtar E.S, Balkaya A. (2010) Production of in vitro haploid plants from in situ induced haploid embryos in winter squash (Cucurbita maxima Duchesne ex Lam.) via irradiated pollen. Plant Cell Tissue Org Cult 102(3):267–277.

17. Metwally E.I., Haroun S.A., El-Fadly G.A. (1996) Interspecific cross between Cucurbita pepo L. and Cucurbita martinezii through in vitro embryo culture. Euphytica 90: 1–7

18. Suprunova T., Shmykova N. In vitro induction of haploid plants in unpollinated ovules, anther and microspore culture of Cucumis sativus // Cucurbitaceae 2008. Proceedings of the IXth EUCARPIA meeting on genetics and breeding of Cucurbitaceae. 21-24 May 2008 Avignon, France. P.371-374.

19. Varghese B.M. (1973) Studies on the cytology and evolution of south Indian Cucurbitaceae. Ph.D. Thesis Kerala Univ., India.

20. Whitaker, T.W. (1933). Cytological and Phylogenetic Studies in the Cucurbitaceae. Botanical Gazette, 94(4), 780–790. doi:10.1086/334347

21. Whitaker T.W., Davis G.N. (1962) Cucurbits. Interscience Pub., Inc.,New York.