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dentification Novel phaC1 Gene Native Pseudomonas putida KT2442 Key Gene for PHA Biosynthesis
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1.3 Sequence analysis of the amplicon from
phaC1
gene
The 540-bp PCR product from native
P. putida
sample
was purified and sequenced. The purification product
is showed in Figure 3.
Figure 2 Gel electerophoresis of PCR production of 540
fragment from high conservative area in
phaC1
Gene
Note: Lane 1 and 2, 50 bp molecular weigh, Lane3-8, 540
fragment PCR product from high conservative sequences in
phaC1
gene in
pseudomonas putida
KT2442, (different
reaction)
Figure 3 The purification of 540 bp PCR fragment from
P.
putida
KT2442
Note: Line 1, 50bp molecular weigh and line 2, 540 bp PCR
product
The 540 bp PCR fragment were purified and
sequenced using the Forward and Revers primer as
demonstrated in Figure 4 and Figure 5 respectively.
A BLASTX search using the selected 190-bp
P. putida
phaC1
gene sequences as query yielded an amino acid
sequence matches with more than 20 PHA synthases
(data not shown). The alignments with the most
homology were those with the PHA synthase 1&2
from
Pseudomonas aeruginosa
PHA synthase 1&2
from
P. putida
etc. Seemingly, the particular bacterial
clone chosen for the sequence analysis contained an
amplified segment of the
Pseudomonas putida phaC1
gene. The sequences in FASTA format were used in
Mega5 application to built the phylogenic tree.
1.4 Phylogenetic tree
The ClustalW method in Mega5 application was used
to make a phylogenic tree. The “Phylip Tree” section
gives the tree in newick format. Figure demonstrated
the phylogenetic tree in newick format (Figure 6).
Figure 6 The phylogenic tree of
phaC1
gene from
pseudomonas putida
KT2442
2 Conclusion
Because of The PHA numerous applications in
industry, medicine and agriculture, the need for
extensive research on genes involved in the
production, it is very much needed. Multiple use of
plastics in the world is increasing day by day, so that
the global market for biodegradable polymers to 114
million pounds, up until 2010, was rape, For this
reason, the production of PHAs are highly regarded
scientists and the significant research on biopolymers
is flow in Europe and Japan and to some extent in
America (European Bioplastics, 2007).
What has attracted great industrial interest is the idea
that application-specific, novel polymers with
physical and chemical properties can be synthesized
merely by controlling the composition of the carbon
source. It is important to note that several economic
and technical barriers to the production of PHAs and
Computational
Molecular Biology