Latest revision as of 02:12, 18 October 2014

TEC-CEM | Parts

ITESM-CEM | Enzy7-K me

Parts 3256

_{Our Parts}

_{List of our parts}

Our Parts

The main goal of our project was to establish the construct which will help us to metabolize 7-ketocholesterol, consisting in the use of three specific enzymes, but for further applications we submitted them in single modules. This will serve as the basis of a future library for standardized work related to atherosclerosis.

Cholesterol Oxidase

This enzyme was first detected in Chromobacterium sp. We introduced it in a plasmid backbone with chloramphenicol resistance: pSB1C3. Its length is of 1871 nucleotides and its codons were optimized in order to use it on E. coli, it already included a stop codon for transcription, it was also modified by the addition of a glycosilation site (NIT) and the peptide signal of human S-cathepsin.

Oxoacyl Reductase

This enzyme was detected in Rhodococcus jostii . We introduced it in a plasmid backbone with chloramphenicol resistance: pSB1C3. Its length is of 1007 nucleotides and its codons were optimized in order to use it on E. coli, it already included a stop codon for transcription, it was also modified by the addition of a glycosilation site (NIT) and the peptide signal of human S-cathepsin.

7-dehydratase

This enzyme (7-alpha dehydratase) was detected in Rhodococcus jostii . We introduced it in a plasmid backbone with chloramphenicol resistance: pSB1C3. Its length is of 602 nucleotides and its codons were optimized in order to use it on E.coli, it already included a stop codon for trancription, it was also modified by the addition of a glycosilation site (NIT) and the peptide signal of human S-cathepsin.

Neomycin Resistance

This selective marker was obtained from an mammalian expression vector. NeoR's length is 855 nucleotides and it was isolated from pcDNA3.1(-)/myc-His A.

BGHPA

Bovine Growth Hormone Polyadenilation Signal for nuclease resistance. Translation terminator for eukaryotic cells.

PCMV

Constitutive promoter from Cytomegalovirus, this promoter works on eukaryotic cells, driving protein expression.

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 <iframe align="right" width="660" height="515" src="//www.youtube.com/embed/1mFwPDbb7UY" frameborder="0" allowfullscreen></iframe>
-<p style="text-align: justify; text-justify: inter-word;"> The main goal of our project was to establish the construct which will help us to degrade 7-ketocholesterol consisting in the use of three specific enzymes, but for further applications we submitted them in single modules. This will serve as the basis of a future library for standardized work related to atherosclerosis.
+<p style="text-align: justify; text-justify: inter-word;"> The main goal of our project was to establish the construct which will help us to metabolize 7-ketocholesterol, consisting in the use of three specific enzymes, but for further applications we submitted them in single modules. This will serve as the basis of a future library for standardized work related to atherosclerosis.
 </p><br>
 <h2>Cholesterol Oxidase</h2>
-<p style="text-align: justify; text-justify: inter-word;"> This enzyme was first detected in <u>Chromobacterium sp.</u> We introduced it in a plasmid backbone with chloramphenicol resistance. Its length is of 1871 nucleotides and its codons were optimized in order to use it on E.coli, it already included a stop codon, it was also modified by the addition of a glycosilation site and the peptide signal of human cathepsin.
+<p style="text-align: justify; text-justify: inter-word;"> This enzyme was first detected in <u>Chromobacterium sp.</u> We introduced it in a plasmid backbone with chloramphenicol resistance: pSB1C3. Its length is of 1871 nucleotides and its codons were optimized in order to use it on <u>E. coli</u>, it already included a stop codon for transcription, it was also modified by the addition of a glycosilation site (NIT) and the peptide signal of human S-cathepsin.
 </p> <br>
 <h2>Oxoacyl Reductase</h2>
-<p style="text-align: justify; text-justify: inter-word;"> This enzyme was detected in Rhodococcus jostii . We introduced it in a plasmid backbone with chloramphenicol resistance. Its length is of 1007 nucleotides and its codons were optimized in order to use it on E.coli, it already included a stop codon, it was also modified by the addition of a glycosilation site and the peptide signal of human cathepsin.
+<p style="text-align: justify; text-justify: inter-word;"> This enzyme was detected in <u>Rhodococcus jostii </u>. We introduced it in a plasmid backbone with chloramphenicol resistance: pSB1C3. Its length is of 1007 nucleotides and its codons were optimized in order to use it on <u>E. coli</u>, it already included a stop codon for transcription, it was also modified by the addition of a glycosilation site (NIT) and the peptide signal of human S-cathepsin.
 </p> <br>
 <h2>7-dehydratase</h2>
-<p style="text-align: justify; text-justify: inter-word;"> This enzyme (7-alpha dehydratase) was detected in Rhodococcus jostii . We introduced it in a plasmid backbone with chloramphenicol resistance. Its length is of 602 nucleotides and its codons were optimized in order to use it on E.coli, it already included a stop codon, it was also modified by the addition of a glycosilation site and the peptide signal of human cathepsin.
+<p style="text-align: justify; text-justify: inter-word;"> This enzyme (7-alpha dehydratase) was detected in <u>Rhodococcus jostii</u> . We introduced it in a plasmid backbone with chloramphenicol resistance: pSB1C3. Its length is of 602 nucleotides and its codons were optimized in order to use it on <u>E.coli</u>, it already included a stop codon for trancription, it was also modified by the addition of a glycosilation site (NIT) and the peptide signal of human S-cathepsin.
 </p> <br>
 <h2>Neomycin Resistance</h2>
-<p style="text-align: justify; text-justify: inter-word;"> This selective marker was gotten from a plasmid for mammalian expression, its length is of 855 nucleotides,and it was isolated from pcDNA3.1(+)/myc-His A.
+<p style="text-align: justify; text-justify: inter-word;"> This selective marker was obtained from an mammalian expression vector. NeoR's length is 855 nucleotides and it was isolated from pcDNA3.1(-)/myc-His A.
 </p><br>
 <h2>BGHPA</h2>
-<p style="text-align: justify; text-justify: inter-word;"> Stop coding for eucaryotic cells
+<p style="text-align: justify; text-justify: inter-word;"> Bovine Growth Hormone Polyadenilation Signal for nuclease resistance. Translation terminator for eukaryotic cells.
 </p><br>
 <h2>PCMV</h2>
-<p style="text-align: justify; text-justify: inter-word;"> Promoter from Human Cytomegalovirus, this promoter works only on eucaryotic cells.
+<p style="text-align: justify; text-justify: inter-word;"> Constitutive promoter from Cytomegalovirus, this promoter works on eukaryotic cells, driving protein expression.
 </p><br><br><br>

Team:ITESM-CEM/Parts

From 2014.igem.org