Fifty plants of 60 day-old cole (Zheyou No. 18) or rice (Ezhong
No. 5) were collected from the crop field rotated by rice and cole.
Five gram of the stems and leaves were cut into pieces (1 cm).
The fragments were surface-sterilized by soaking in 75% ethanol
for 1 min, and in 0.1% mercuric chloride (leaves for 2 min, stems
for 4 min). They were washed several times in sterile water,
transferred to an autoclaved mortar, and homogenized with
a mixture of quartz sand and sterile water. An aliquot (100 ml) of
the homogeneous suspension was spread on sterile potato
dextrose agar (PDA), and incubated at 27 C. After incubation for
48 h, single colonies were picked up and tested their inhibiting
activity against fungal pathogens. Among seven endophytic
isolates, only strain CHM1 significantly (p < 0.01) inhibited
growth of six species of fungal pathogens in vitro, as mentioned
in Section 2.1.
The culture solution of CHM1 was prepared by shaking LB liquid
culture at 30 C, 170 rpm for 36 h. It was further centrifuged at
6000 g for 5 min at 4 C and the supernatant collected and
passed through a 0.22-mm filter, yielding CHM1 culture filtrate. The
sterile culture filtrate was obtained by autoclaving the resultant
culture filtrate at 121 C for 30 min