Difference between revisions of "Part:BBa K2365039"

Latest revision as of 06:00, 11 October 2022

ADH1 promoter

AHD1 Yeast promoter Constitutive expression: Alcohol dehydrogenase I promoter, full length version is very strong which can promote high expression. Truncated promoters are constitutive and have low expression. Widely exist in human and animal liver, plant and microbial cells as a key enzyme in short-chain alcohol metabolism. Glucose can promote the presence of expression, in many physiological processes play an important role.

Sequence and Features

Assembly Compatibility:

10
COMPATIBLE WITH RFC[10]
12
COMPATIBLE WITH RFC[12]
21
COMPATIBLE WITH RFC[21]
23
COMPATIBLE WITH RFC[23]
25
COMPATIBLE WITH RFC[25]
1000
COMPATIBLE WITH RFC[1000]

Improved design:

We construct the device：ADH1 promoter+GFP+CYC1 terminator.ADH1 promoter is a truncated promoter, GFP is a mutant. These is a invariable sequence upstream of GFP coding sequence.

The promoter of yeast alcohol dehydrogenase (ADH1) is widely used for the expression of heterologous genes in Saccharomyces cerevisiae. We tested the expression of GFP proteins in five transformants containing the ADH1 promoter. Our data shows that there are some differences in the fluorescence intensity of different yeast transformants with ADH1 promoter. The variation trends of promoter strengths from early-log to stationary phase varied in different transformants. Individual transformants colonies have different expression efficiency. We will test this promoter for more than three times.

Improved part by UTokyo 2022:

We created Improved part BBa_K4238000 based on part BBa_K2365039. We attempted to create a shorter, and therefore easier to use for genetic engineering, and more powerful basic part of a promoter to make more options for easy-to-handle constitutive promoters. ADH1 is alcohol dehydrogenase I (ADH1), and its promoter of yeast, especially the medium-length one, is often used as a constitutive expression promoter for yeasts. The part called BBa_K2365039 is a medium-length ADH1 promoter whose length is 618 bp. We developed a promoter whose length is 409 bp by deleting the upstream 209 bp of this part.
The upstream sequence we deleted contained an activation sequence called UASrpg (upstream activation sequence of ribosome protein genes), so we could expect an increase in the expression level.[1]

Figure1. Distribution of fluorescence. 257 cells of ADH1p long and 418 cells of ADH1p short were observed.

Figure2. Average of fluorescence. Error bars indicate standard errors. * denotes a significant difference at $p<0.05$, ** denotes a significant difference at $p<0.01$, and n.s. denotes no significant difference. The statistical test was conducted using the Wilcoxon rank sum test.

The expression of the fluorescent protein increased by using ADH1p short.

@@ Line 3: / Line 3: @@
 <partinfo>BBa_K2365039 short</partinfo>
-AHD1 Yeast promoter Constitutive expression
+AHD1 Yeast promoter Constitutive expression:
+Alcohol dehydrogenase I  promoter, full length version is very strong which can promote high expression. Truncated promoters are constitutive and have low expression. Widely exist in human and animal liver, plant and microbial cells as a key enzyme in short-chain alcohol metabolism. Glucose can promote the presence of expression, in many physiological processes play an important role.
 <!-- Add more about the biology of this part here
@@ Line 15: / Line 15: @@
 ===Improved design:===
+[[File:2HXT7p.jpeg|300px|center]]
 We construct the device：ADH1 promoter+GFP+CYC1 terminator.ADH1 promoter is a truncated promoter, GFP is a mutant. These is a invariable sequence upstream of GFP coding sequence.
 <!-- -->
 ===[[File:图片1.jpg|500px|center]]===
 The promoter of yeast alcohol dehydrogenase (ADH1) is widely used for the expression of heterologous genes in Saccharomyces cerevisiae. We tested the expression of GFP proteins in five transformants containing the ADH1 promoter. Our data shows that there are some differences in the fluorescence intensity of different yeast transformants with ADH1 promoter. The variation trends of promoter strengths from early-log to stationary phase varied in different transformants. Individual transformants colonies have different expression efficiency. We will test this promoter for more than three times.
+===Improved part by UTokyo 2022:===
+<html>
+We created Improved part <a href="https://parts.igem.org/Part:BBa_K4238000">BBa_K4238000</a> based on part <a href="https://parts.igem.org/Part:BBa_K2365039">BBa_K2365039</a>.
+We attempted to create a shorter, and therefore easier to use for genetic engineering, and more powerful basic part of a promoter to make more options for easy-to-handle constitutive promoters. ADH1 is alcohol dehydrogenase I (ADH1), and its promoter of yeast, especially the medium-length one, is often used as a constitutive expression promoter for yeasts. The part called <a href="https://parts.igem.org/Part:BBa_K2365039">BBa_K2365039</a> is a medium-length ADH1 promoter whose length is 618 bp. We developed a promoter whose length is 409 bp by deleting the upstream 209 bp of this part.<br />
+The upstream sequence we deleted contained an activation sequence called UASrpg (upstream activation sequence of ribosome protein genes), so we could expect an increase in the expression level.[1]<br />
+</html>
+[[File:ADH1ps4.png|550px|thumb|center|Figure1. Distribution of fluorescence. 257 cells of ADH1p long and 418 cells of ADH1p short were observed.]]
+[[File:ADH1ps5.png|550px|thumb|center|Figure2. Average of fluorescence. Error bars indicate standard errors. * denotes a significant difference at $p<0.05$, ** denotes a significant difference at $p<0.01$, and n.s. denotes no significant difference. The statistical test was conducted using the Wilcoxon rank sum test.]]
+<html>
+The expression of the fluorescent protein increased by using ADH1p short.
+<br />
+</html>