In-situ Detection Of Carotenoid Biosynthetic Gene Expression During Cara Cara Navel Orange (citrus Sinensis Osbeck) Fruit Development

Citrus fruit is reported to be carotenoid abundant,normally more carotenoids content was found in the peel than that in the pulp.Some red-fleshed mutant such as 'Cara Cara' navel orange with abnormal lycopene accumulation and attracting appearance are reported to be good research material not only for the elucidation of bud sport mechanism in citrus fruits but also for the reveal of carotenoid biosynthetic pathway in citrus.In the present research,the red-fleshed 'Cara Cara' navel orange(C. sinenesis Osbeck) and the normal orange-fleshed "Seika" navel orange(C.Sinenesis Osbeck) fruit were used to investigate the in situ mRNA accumulating of carotenoid biosynthesis related genes on the cellular level at different ripening stage.Peel and pulp color development in 'Cara Cara' and "Seika" navel orange was detected with chromameter.The main results are as follows:1.A special red thread-like structure in 'Cara Cara' juice sac was observed with hand-section preparation and fixation treatment under microscope observation.The structure is the same as the plastid structure in lycopene-abundant crop tomato(Lycopersicon esculentum Miller) and also visible in the red pummelo(Citrus grandis L.Osbeck.) pulp.The increasing density of the structure in developing 'Cara Cara' juice sac is correlated with the lycopene accumulation. 2.Paraffin-embed tissue section of flavedo,albedo,segment and juice sac dissected from 'Cara Cara' and 'Seika' navel orange fruits at different stages were prepared for the RNA in-situ hybridization,using the cRNA listed above,the accumulation of psy,pds,lcyb mRNA was visible at the epidermal cell under cortex in the flavedo part of 'Cara Cara' and 'Seika' navel orange,the expression level of such genes were different depending on the fruit maturing process.Seldom mRNA accumulation could be detected at the ocellus or the periphery cells in the flavedo. The expression profile in the albedo and juice sac cells was distance from the corresponding ceils in the flavedo,to our surprise,the mRNA accumulation in the vascular bundle was negligible.And generally,the mRNA expression in the albedo, segment and juice sac was lower than that in the flavedo.