Abstract:

In this study，transient expression system was used to analyze the function of
β-ketocarotase for biosynthesis of ketocarotenoid in pumpkin fruit. The Agrobacteria that contained
pBI121-CMTPCRBKT vector with CrBKT gene and the control pBI121 vector were individually injected
into the pumpkin fruits at 2，5，10，15 and 25 days after pollination，respectively. It was found that the fruit
pulp was light reddish in 2- and 5- day fruits with darker color in pulp of 5-day fruit while there was no red pigment accumulated in more than 10-day fruit. High performance liquid chromatography（HPLC）
analysis of pigment composition revealed that the accumulated red pigments were canthaxanthin and
astaxanthin. Compared with the control，the carotenoid content in the 2- and 5-day fruits injected with the
CrBKT gene were significantly increased，which increased by about 1/3 in 5-day young fruit and
ketocarotenoid content was 106.31 μg · g-1，of which canthaxanthin accounted for 80.65%，astaxanthin was
19.35%，while the carotenoid content of the fruits in more than 10-day had no obvious change and there
was no accumulation of ketocarotenoid. PCR amplification indicated that CrBKT was expressed in the
transformed tissue. The result showed that the CrBKT gene could be only expressed in less than 5-day fruit
and it could convert carotenoid in young pumpkin into ketocarotenoid. However，the expression of
exogenous gene was discouraged due to the bacteria could not osmotic into the flesh tissue with the fruit
ripening.

Key words: pumpkin； Chlamydomonas reinhardtii；&beta, -carotene ketolase；gene；transient expression