全 文 ：浙江大学学报 (农业与生命科学版 )　 26( 5): 579～ 580, 2000
Journal of Zhejiang Un iversity ( Ag ric. & Life Sci. )
　　Received date: 2000-03-08
Foundation item: The National Natural Science Foundation of China ( 39830230)
Biography: LI Ying, female, born in 1973 in Xintai , Sh andong , Ph. D. s tud ent , s peciali zed in v egetable ph ysiology and bio-
chemis try.
Article ID: 1008-9209( 2000) 05-0579-02
SDS-PAGE Analysis of Protein Characteristics in
Developing Fruit of Lagenaria leucantha
LI Ying , YU Jing-quan, ZHU Zhu-jun
(Dept. of Horticulture, Zhejiang Univ. , Hangz hou 310029, China)
瓠瓜果实发育中的蛋白质 SDS-PAGE分析
李 英 ,喻景权 ,朱祝军
(浙江大学 园艺系 ,浙江杭州 310029)
　　 In seed plants, successful frui t set and subsequent development are dependent on po llination
and fer ti li zation since fertilization activ ates cell division and trig gers f rui t development by produc-
ing phytohormones especially auxins and GAs. Fruit set was very low in Lagenaria leucantha
Rusby during ea rly spring. We found that CPPU [N-( chloro-4-py ridyl ) -N′-phenyurea ], a new
kind of cy tokinin, was mo re ef fective than N A A and GA3 in inducing pa rthenocarpic f rui t g row th
and in decreasing f ruit abor tion. In this paper , w e examined the possible role o f ferti li zation and
CPPU in protein synthesis in developing f rui t of Lagenaria leucantha by SDS-PAGE analy sis.
1　Materials and Methods
1. 1　 Plant materials　　 Seeds o f Lagenaria leucantha Rusby ( cv. Hang zhou long-gourd) w ere
sow n on 15 February 1999, and the seedling s were t ransplanted on 20 March. There w ere three
t reatments: Nonpol lination; Hand pollination, and Nonpo llina tion+ 50 mg /L CPPU. Unpol linat-
ed ovaries w ere prepa red by bagging the female flowers one day befo re anthesis. The solution of
CPPU was applied wi th a spray to ovaries of bagged female flow ers at anthesis a t the rate of 0. 5
m L per ovary. Frui t leng th w as recorded every three days. Fo r assay s o f pro tein, the f lesh of
f rui ts a t anthesis and 8 days af ter t reatment wa s f ro zen in liquid ni t rog en and sto red a t - 20℃ .
1. 2　 Ex traction o f pro teins　　 Half g ram of f ruit ti ssue w as homogenized in 1 mL ex tract buffer
containing 25 mmol /L DTT and 250 mmol /L Tris-HCl, pH6. 8. The homogenate w as centri fuged
a t 12 500 g for 20 min. Aliquo t o f the 250μL supernatant was added into an eppendo rf tube filled
w ith 1 mL of 80% acetone, then f ro zen in - 20℃ for 10 min. Af ter thaw ing at 25℃ , i t w as
centrifug ed at 10 000 g fo r 10 min, the supernatant was ca refully discarded.
1. 3　 SDS-PAGE　　 Pro tein ex t racts were resuspended in SDS buffer containing 50 mmol /L
Tris-HCl ( pH 6. 8) , 10% g lycero l, 2% SDS, 0. 1% bromophenol and 5%β - mercaptoethanol.
Fig. 1　 Ef fects of nonpollination, pollination
and CPPU on the length of fruits in
Lagenaria leucantha
After heating in a boi ling w ater bath for 3 min, 5μL
of ex tracts w as loaded onto a 12% SDS gel. Elect ro-
po retic separation o f pro tein w as conducted a t 25℃
using a constant current of 30 mA. Pro tein bands in
the gel w ere stained wi th Coomassie Bri llia nt Blue.
2　Results and Discussion
2. 1　 CPPU induction of par thenocarpic f rui t　　 Un-
pollinated ova ries did no t develop and the f rui t leng th
remained almost constant. How ever, po llination
great ly improved the g row th o f ovaries and frui t
leng th reached near 40 cm in 12 days af ter anthesis.
CPPU showed high activi ty in inducing pa rtheno-
carpic f ruit g row th o f unpollinated ovary. Fur ther-
mo re the CPPU treated f rui ts g rew much faster than the pollinated f rui ts ( Fig 1) .
Fig. 2　 SDS-PAGE prof iles of proteins f rom
fruits at anthesis 1 and 8 days after
treatment ( nonpollination 2, pollina-
tion 3 and Nonpollination+ CPPU
treatment 4)
2. 2　 SDS-PAGE analysis　　 SDS-PAGE analysis o f
to tal so luble pro tein in ovaries rev ealed that there
w ere no significant di fferences in the frui t pro tein
profi le among ova ries at anthesis, pollina ted and CP-
PU treated ovaries, and most of them are of quanti ta-
tiv e nature ( Fig 2) . Some proteins ( 36. 1 kD, 37. 2
kD, 74. 7 kD) , how ever, disappea red in unpollina ted
f rui t. It seemed likely that pollination and CPPU
treatment improve f rui t development mainly by stim-
ulating new synthesis o f tho se pro teins tha t already
existed or slowing down thei r deg rada tion. It is also
possible that pollinated and CPPU-treated f rui ts pro-
duced some new proteins wi th molecular w eigh ts sim-
ilar to the pro teins w hich already ex isted, or the new
pro teins w ere too li t tle to be detected by SDS-PAGE.
Unpollinated f rui t abo rted due to the lack of this abi li ty.
(责任编辑　邓君奇 )　　
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