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Legume Genomics and Genetics (online), 2010, Vol. 1, No.3, 11-17
http://lgg.sophiapublisher.com
Figure 1 Alignment of the complete deduced amino acid sequences of β-ketoacyl-ACP synthase
Ⅰ
genes
strain names. The four conserved
Note: Conserved amino acid residues are highlighted in black boxes. The sequences are denoted by their
residues of
A. hypogaea
KAS
Ⅰ
corresponding to
E. coli
FabB Cys163, His298, Lys328 and His333 are highlighted in asterisks. Accession
numbers for the sequences were as follows:
Arachis hypogaea
(FJ768729),
Glycine max
(AAF61730),
Arabidopsis thaliana
(AAC49118),
Ricinus communis
(AAA33873),
Escherichia coli
(AAC67304),
Jatropha curcas
(ABJ90468),
Helianthus annuus
(ABM53471)
Figure 2 Neighbor-joining tree based on the deduced amino
ccession numbers and strain
f
AhKAS
tative real-time PCR (qRT-PCR) was employed
acid sequences of
KAS
Ⅰ
homologs
Note: Sequences are shown by their a
names; Bootstrap values from neighbor-joining analyses are
listed to the left of each node, with values more than 50 are shown
pET vector system in
E. coli
. Gene expression and the
molecular weight of this enzyme were monitored by
SDS-PAGE (Figure 3). After IPTG induction, a novel
polypeptide with the molecular mass of approximately
49 kD was expressed in
E. coli
as revealed by SDS-
PAGE. The recombinant
protein reached maximal
expression levels 4 to 8 h after induction and showed
a similar molecular weight of approximately 49 kD
compared to the deduced value.
1.4 Quantitative real-time PCR analysis o
Ⅰ
gene
The quanti
to confirm the expression patterns of
AhKAS
Ⅰ
gene
in four peanut tissues and at different developmental
stages of seeds. β-actin was used as an internal refe-
rence control for total RNA input. β-actin PCR pro-
duct was not detected when reverse transcriptase was
omitted, indicating that the RNA template was free of
genomic DNA. The results revealed that
AhKAS
Ⅰ
gene displayed tissue-specific expression patterns in
peanut (Figure 4). The
AhKAS
Ⅰ
gene was expressed
Figure 3 The SDS-PAGE map of
AhKAS
Ⅰ
protein pression
ex
Note: M: Protein marker MP102 (Tiangen, China); 1: AhKAS
Ⅰ
recombination protein containing His.Tag not induced with
IPTG; 2~6: AhKAS
Ⅰ
recombination protein containing His.
Tag induced with IPTG for 1 h, 2 h, 4 h, 6 h and 8 h, respectively
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