Difference between revisions of "Part:BBa K3971030"

Latest revision as of 22:23, 21 October 2021

Cassette to insert spp gene in the Neutral Site 3 of the S. elongatus UTEX 2973 genome.

spp codes for Sucrose-phosphate phosphatase (SPP), an enzyme involved in the sucrose biosynthesis pathway in the cyanobacteria Synechocystis PCC 6803 [1].

This composite part has been designed to insert the spp gene into the Neutral Site 3 of the S. elongatus UTEX 2973 genome.

The promoter in this part is PcpcB-m6 (BBa_K3971000). In order to allow for modularity in the promoter region of the cassette, two restriction sites Nde1 and BamH1 flank it, so that the promoter can be easily excised out and replaced with another. Thus one can study the effect of different promoters on sucrose production efficiency.

Both ends of the composite part contain Kpn1 restriction sites since this cassette is designed to be cloned into the pSL2680 plasmid, which is a Cpf1 based CRISPR plasmid [2], which has a Kpn1 site and a Sal1 site to allow for cloning of cassettes.

We aimed to overexpress the spp gene in order to increase the amount of sucrose produced by S. elongatus when exposed to salt stress.

Figure of the composite construct for inserting the spp gene into the Neutral Site 3 of the S. elongatus UTEX 2973 genome. NS3a and NS3b are the two homology arms to Neutral Site 3.

References

1:Lin, Po-Cheng, Fuzhong Zhang, and Himadri B. Pakrasi. "Enhanced production of sucrose in the fast-growing cyanobacterium Synechococcus elongatus UTEX 2973." Scientific reports 10.1 (2020): 1-8.

2: Du, Wei, et al. "Exploring the photosynthetic production capacity of sucrose by cyanobacteria." Metabolic engineering 19 (2013): 17-25.

Sequence and Features

Assembly Compatibility:

10
INCOMPATIBLE WITH RFC[10]
Illegal PstI site found at 635
Illegal PstI site found at 2220
12
INCOMPATIBLE WITH RFC[12]
Illegal PstI site found at 635
Illegal PstI site found at 2220
21
COMPATIBLE WITH RFC[21]
23
INCOMPATIBLE WITH RFC[23]
Illegal PstI site found at 635
Illegal PstI site found at 2220
25
INCOMPATIBLE WITH RFC[25]
Illegal PstI site found at 635
Illegal PstI site found at 2220
1000
COMPATIBLE WITH RFC[1000]

@@ Line 3: / Line 3: @@
 <partinfo>BBa_K3971030 short</partinfo>
-This composite part has been designed to insert the spp gene into the Neutral Site 3 of the S. elongatus UTEX 2973 genome.
+''spp'' codes for Sucrose-phosphate phosphatase (SPP), an enzyme involved in the sucrose biosynthesis pathway in the cyanobacteria ''Synechocystis'' PCC 6803 [1].
+This composite part has been designed to insert the ''spp'' gene into the Neutral Site 3 of the ''S. elongatus'' UTEX 2973 genome.
 The promoter in this part is PcpcB-m6 (BBa_K3971000). In order to allow for modularity in the promoter region of the cassette, two restriction sites Nde1 and BamH1 flank it, so that the promoter can be easily excised out and replaced with another. Thus one can study the effect of different promoters on sucrose production efficiency.
-Both ends of the composite part contain Kpn1 restriction sites since this cassette is designed to be cloned into the pSL2680 plasmid, which is a Cpf1 based CRISPR plasmid [1], which has a Kpn1 site and a Sal1 site to allow for cloning of cassettes.
+Both ends of the composite part contain Kpn1 restriction sites since this cassette is designed to be cloned into the pSL2680 plasmid, which is a Cpf1 based CRISPR plasmid [2], which has a Kpn1 site and a Sal1 site to allow for cloning of cassettes.
@@ Line 16: / Line 18: @@
 ===References===
-: Du, Wei, et al. "Exploring the photosynthetic production capacity of sucrose by cyanobacteria." Metabolic engineering 19 (2013): 17-25.
+:Lin, Po-Cheng, Fuzhong Zhang, and Himadri B. Pakrasi. "Enhanced production of sucrose in the fast-growing cyanobacterium Synechococcus elongatus UTEX 2973." Scientific reports 10.1 (2020): 1-8.
+: Du, Wei, et al. "Exploring the photosynthetic production capacity of sucrose by cyanobacteria." Metabolic engineering 19 (2013): 17-25.