Figure Legend Fig. S1.
description
Transcript of Figure Legend Fig. S1.
Heterologous expression system in Aspergillus oryzae for fungal biosynthetic gene clusters of secondary metabolites
Applied Microbiological Biotechnology Kanae Sakai,1 Hiroshi Kinoshita,1 and Takuya Nihira1,2*
1International Center for Biotechnology, Osaka University, 2-1 Yamadaoka, Suita, Osaka 565-0871, Japan2MU-OU Collaborative Research Center for Bioscience and Biotechnology, Faculty of Science, Mahidol University, Rama VI Rd., 10400 Bangkok, Thailand
*Corresponding author. Tel.: +81-6-6879-7452; Fax: +81-6-6879-7454; E-mail: [email protected]
Figure Legend
Fig. S1.
Representative genotype of transformants. (a) Southern blot analysis of the MK transformants. Southern blot analysis was carried
out against MluI-digested genomic DNA using the mokB fragment as the probe. The mokB fragment was amplified by PCR with
primers mokB-f and mokB-r. Lanes C1, cosmid 12-33; C2, cosmid sCnDmokB; W, A. oryzae NS4 strain; M, λ-EcoT14I digest DNA
marker; 1, M12-33-5; 2 to 4, candidate transformants (MK5-1,3,4); 5, M12-33-17; 6 to 9, candidate transformants (MK17-1, 2, 5,
6); 10, M12-33-18; 11, candidate transformants (MK18-1). The bands indicated by arrow were derived from cosmid 12-33. The
band indicated by arrowhead was derived from sCnDmokB. (b) Southern blot analysis of the MK-L transformants. Southern blot
analysis was carried out against SnaBI-digested genomic DNA using the AolaeA PCR fragment (amplified by AolaeA-3f: 5’-
atggcctaatgtacgccc-3’, AolaeA-3r: 5’-ctcctgcagagtctcggat-3’) as the probe. Lanes V, pPTRI-laeA; M, λ-EcoT14I digest DNA
marker; 1, M12-33-17 strain; 2, MK-L1; 3, MK-L2.
The band indicated by arrow was derived from pPTRI-laeA. The band indicated by arrowhead was the signal of native laeA gene.
(c) Southern blot analysis of KsMK transformants. Southern blot analysis was carried out against EcoRI-digested genomic DNA
using the mokA or mokB2 PCR fragment (amplified by mokA-f, -r or mokB-2f, -2r primer sets) as the probe. Lanes: H, A. oryzae
host strain (Ks31); 6, 21, candidate transformants (KsMK-6, 21). (d) Southern blot analysis of TQ transformants. Southern blot
analysis was carried out against KpnI-digested genomic DNA using the tdiD PCR fragment as a probe. Lanes: V, TQ gene cluster
expression vector sCsCTQ; M, λ-EcoT14 digest DNA marker; H, Ks31 host; 25, 33, 34,36, 39, 47, 52, 53, candidate transformants
(KsTQ-25, 33, 34, 36, 39, 47, 52, 53). The arrowhead indicated the position of 9.4 kb.
FIG. S2. 1H-NMR spectrum of MK purified A. oryzae transformant KsMK-25
FIG. S3. 1H-NMR spectrum of MJ purified A. oryzae transformant KsMK-25
FIG. S4. 1H-NMR spectrum of TQ purified A. oryzae transformant KsTQ-39
FIG. S1
mokA
mokB2
(probe)
c6H 21
a
b MV 1 2 3
C1 C2 WM 1 2 3 4 5 6 7 8 9 10 11
V 25 33 34 36 39 47 52 53H
d
FIG. S2
FIG. S3
FIG. S4