The Study Of Karyotype On Genus Tagetes L. And Factors In The Genetic Transformation System Of Psy Gene For Tagetes Erecta L.

Karyotypes of eight genus Tagetes L. accessions were studied, and he result could provide the scientific basis for genetic breeding in the cytology level. Based on the marigold (Tagetes erecta L.) N-091, the N-092, N-093, the N-094 for the materials, according to transient expression of gus gene, the suitable kind of variety and explant types for conversion have been chosen, and the factors in the Agrobacterium-mediated genetic transformation system have been studied, This will provide basic information for new marigold which with higher lutein content by making psy gene overexpressied. The detailed study results are as follows.(1) The karyotype formula of T. erecta L. Harvest , Taishan , Marval , Perfection Yellow , Perfection Orange , Inca Orange , Inca Yellow was like: 2n=2x=6sm+18m,2n=2x=14sm+10m,2n=2x=24m,2n=2x=4sm+20m,2n=2x=24m, 2n=2x=2sm+22m,2n=2x=24m; the length type was 2n=4L+6M₂+10M₁+4S, 2n=6L+4M₂+10M₁+4S,2n=4L+8M₂+8M₁+4S,2n=6L+6M₂+6M₁+6S,2n=2x=4sm+20m,2n=4L+8M₂+8M₁+4S,2n=4L+4M₂+12M₁+2S;The KA type was of 1B, 2B, 1B, 1B, 1B, 1B, 1B. The karyotype formula of T.patula L. Golden Gate was 2n=4x=48=4sm+44m; the length type was 2n=12L+8M₂+16M₁+12S; The KA type was of 1B.(2) Based on the marigold N-091, the N-092, N-093, the N-094 leaves and stems with axillary buds for experimental material. Results of gus gene transient expression: N-094 (86.6%) > N-093 (73.3%) > N-091 (43.3%) > N-092 (40%). N-094 was the variety which was most easily converted.(3) For N-094, based on leaves and stems with axillary buds as the explants, the results of regeneration induction rate: stems with axillary buds (93.3%) > leaves (50%). Stems with axillary buds was available to be the explant in the regeneration system of marigold.The inductive medium for stems with axillary buds: MS + 6-BA 0.1 mg/L + NAA 0.1 mg/L + 6 g/L plant gel.(4) According to transient expression of gus gene, the optimal transformation conditions were obtained: take the stems with axillary buds as explants, precultured for 1d, agrobacterium tumefacien infection with OD600=0.8, suspension cultured in1/2 MS0 medium, As 100μmol/L in 1/2 MS0 medium and cocultivation medium, needle puncturing method was more useful during invasion, and co-cultivation for 2 days. In this way a total of 485 explant has been infected, and four resistant plant were got. After the PCR detection, two resistance plants were positive.The rate of positive plant for PCR detection was 0.41%.