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− | <H3> | + | <li class="item"> |
− | Bronze Medal Criterion #4
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| + | <!--内容--> |
| + | <div class="content-middle-outer"> |
| + | <div class="content-middle"> |
| + | <section class="article p-t-30 p-b-54"> |
| + | <h1 class="content-header">Contribution</h1> |
| + | |
| + | <section> |
| + | <h1 class="title">BBa_K4002002</h1> |
| + | |
| + | <h2 class="title2">== Profile BBa_K4002002==</h2> |
| + | <p>Name: HXK1</p> |
| + | <p>Base Pairs: 1458 bp</p> |
| + | <p>Origin: <i>Saccharomyces cerevisiae</i>, genome</p> |
| + | <p>Properties: An enzyme catalyze the phosphorylation of hexose</p> |
| + | |
| + | <h2 class="title2">== Usage and Biology ==</h2> |
| + | <p>This is a sequence coding enzyme HXK1. This enzyme can catalyze the phosphorylation of hexose to |
| + | hexose 6-phosphate (D-glucose 6-phosphate and D-fructose 6-phosphate, respectively). Besides, it can |
| + | mediate the initial step of glycolysis by catalyzing phosphorylation of D-glucose to D-glucose |
| + | 6-phosphate.</p> |
| + | </section> |
| + | |
| + | <section> |
| + | <h1 class="title">BBa_K4002003</h1> |
| + | |
| + | <h2 class="title2">== Profile ==</h2> |
| + | <p>Name: pYES2-gRNA-hyg-MCS</p> |
| + | <p>Base Pairs: 388 bp</p> |
| + | <p>Origin: <i>From article, Addgene</i></p> |
| + | <p>Properties: A piece of RNA complementary to the target gene.</p> |
| + | |
| + | <h2 class="title2">== Usage and Biology ==</h2> |
| + | <p>BBa_K4002003 is a piece of RNAs that function as guides for RNA- or DNA-targeting enzymes, which they |
| + | form complexes with. And this sequence is inserted into plasmid vector.</p> |
| + | |
| + | <h2 class="title2">== Construct design ==</h2> |
| + | <p>sgRNA is inserted into plasmid vector. Plasmid sequence map is shown in Figure 1. </p> |
| + | <div class="img-container m-t-12 m-b-12"> |
| + | <img src="https://static.igem.org/mediawiki/2021/7/74/T--Xiamen_City--img_contribution_1.jpg" alt="" style="width: 80%;"> |
| + | <span class="figure">Figure 1. Schematic map of pYES2-gRNA-hyg-MCS plasmid.</span> |
| + | </div> |
| + | </section> |
| + | |
| + | <section> |
| + | <h1 class="title">BBa_K4002004</h1> |
| + | |
| + | <h2 class="title2">== Profile ==</h2> |
| + | <p>Name: pHCas9-Nours</p> |
| + | <p>Base Pairs: 4837bp</p> |
| + | <p>Origin: <i>Streptococcus pyogenes</i>, Addgene</p> |
| + | <p>Properties: An endonuclease enzyme associated with the CRISPR.</p> |
| + | |
| + | <h2 class="title2">== Usage and Biology ==</h2> |
| + | <p>This is a sequence coding pHCas9 protein. This protein is a dual RNA-guided DNA endonuclease enzyme |
| + | associated with the (CRISPR) adaptive immune system. The Cas9 protein has been heavily utilized as a |
| + | genome engineering tool to induce site-directed double-strand breaks in DNA. The genes that encode |
| + | the Cas9 protein and sgRNA were introduced into a cell and programmed to change its target gene. |
| + | sgRNA has regions that are complementary to the target sequence. A complex consisting of sgRNA and |
| + | Cas9 protein is formed inside the cell and binds to target sites.</p> |
| + | </section> |
| + | |
| + | <section> |
| + | <h1 class="title">BBa_K4002005</h1> |
| + | |
| + | <h2 class="title2">== Profile ==</h2> |
| + | <p>Name: <i>endo-pgaA</i></p> |
| + | <p>Base Pairs: 1113bp</p> |
| + | <p>Origin: <i>Aspergillus niger</i> SC323, genome</p> |
| + | <p>Properties: An enzyme degradation of pectin</p> |
| + | |
| + | <h2 class="title2">==== Usage and Biology ====</h2> |
| + | <p>Polygalacturonase is an enzyme that hydrolyzes the alpha-1,4 glycosidic bonds between galacturonic |
| + | acid residues. It is also known as pectin depolymerase, PG, pectolase, pectin hydrolase, and |
| + | poly-alpha-1,4-galacturonide glycanohydrolase. This part as a repair template DNA was connected with |
| + | homology arm of HXK1.</p> |
| + | </section> |
| + | |
| + | <section> |
| + | <h1 class="title">BBa_K4002006</h1> |
| + | |
| + | <h2 class="title2">== Profile ==</h2> |
| + | <p>Name: HR-L-endo-pgaA-HR-R</p> |
| + | <p>Base Pairs: 2135 bp</p> |
| + | <p>Origin: Ssynthetic</p> |
| + | <p>Properties: CRISPR technology repair template for build a type of multi-functional yeast</p> |
| + | |
| + | <h2 class="title2">== Usage and Biology ==</h2> |
| + | <p>Polygalacturonase is an enzyme that hydrolyzes the alpha-1,4 glycosidic bonds between galacturonic |
| + | acid residues. It is also known as pectin depolymerase, PG, pectolase, pectin hydrolase, and |
| + | poly-alpha-1,4-galacturonide glycanohydrolase. Pectin is a significant carbohydrate component that |
| + | comprises plant cell walls. The brewer's yeast uses sugar in the fruit juice to produce alcohol, and |
| + | pectinase destroys the pectin located in the cell wall to improve the juice yield and eliminate the |
| + | cloudiness of the fruit wine. HR-L and HR-R are homology arm, refers to the flanking sequence on |
| + | both sides of the HXK1.</p> |
| + | |
| + | <h2 class="title2">== Construct design ==</h2> |
| + | <p>HR-L is the homology arm upstream HXK1. HR-R is the homology arm downstream HXK1. pgaA is the |
| + | sequence of pgaA inserted in the homology arm (Figure 1 and 2). </p> |
| + | |
| + | <div class="img-container m-t-12 m-b-12"> |
| + | <img src="https://static.igem.org/mediawiki/2021/2/24/T--Xiamen_City--img_contribution_2.jpg" alt="" style="width: 90%;"> |
| + | <span class="figure">Figure 1. HR-L-endo-pgaA-HR-R box.</span> |
| + | </div> |
| + | |
| + | <div class="img-container m-t-12 m-b-12"> |
| + | <img src="https://static.igem.org/mediawiki/2021/8/87/T--Xiamen_City--img_contribution_3.jpg" alt="" style="width: 90%;"> |
| + | <span class="figure">Figure 2. Schematic map of HR-L-endo-pgaA-HR-R.</span> |
| + | </div> |
| + | </section> |
| + | |
| + | <section> |
| + | <h1 class="title">BBa_K4002007</h1> |
| + | |
| + | <h2 class="title2">== Profile ==</h2> |
| + | <p>Name: <i>Cas9+gRNA+HR-L-endo-pgaA-HR-R</i></p> |
| + | <p>Base Pairs: 7360 bp</p> |
| + | <p>Origin: Synthetic</p> |
| + | <p>Properties: CRISPR technology build a type of multi-functional yeast</p> |
| + | |
| + | <h2 class="title2">== Usage and Biology ==</h2> |
| + | <p>Saccharomyces cerevisiae is a species of yeast (single-celled fungus microorganisms). The species has |
| + | been instrumental in winemaking, baking, and brewing since ancient times. In fruit wine production, |
| + | <i>Saccharomyces cerevisiae</i> uses the sugars in fruit juice to ferment to produce alcohol. It is |
| + | necessary to add pectinase to destroy the pectin in the cell wall to increase the juice yield and |
| + | increase the dissolution of aromatic substances such as pigments or terpenes.</p> |
| + | <p>CRISPRs (Clustered Regularly Interspaced Short Palindromic Repeats) are specific regions in some |
| + | bacterial and archaeal genomes that, together with associated Cas (CRISPR-associated) genes, |
| + | function as an adaptive immune system in prokaryotes. While the specific ‘adaptive’ nature of this |
| + | immunity is still under investigation, it is known that exogenous DNA is processed by Cas proteins |
| + | into short (~30 base pair) sequences that are adjacent to the Protospacer Adjacent Motif (PAM) site. |
| + | These short pieces of DNA are then incorporated into the host genome between repeat sequences to |
| + | form spacer elements. The repeat-spacer-repeat array is constitutively expressed (pre-CRISPR RNAs or |
| + | pre-crRNAs) and processed by Cas proteins to form small RNAs (crRNAs). The small RNAs are then |
| + | loaded into Cas proteins and act to guide them to initiate the sequence-specific cleavage of the |
| + | target sequence.</p> |
| + | |
| + | <div class="img-container m-t-12 m-b-12"> |
| + | <img src="https://static.igem.org/mediawiki/2021/3/37/T--Xiamen_City--img_contribution_4.jpg" alt="" style="width: 80%;"> |
| + | <span class="figure">Figure 1 The principle of CRISPR Cas9.</span> |
| + | </div> |
| + | |
| + | <h2 class="title2">== Construct design ==</h2> |
| + | <p>pHCas9 is a sequence coding endonuclease enzyme associated with the CRISPR. sgRNA is a sequence of |
| + | RNA participant in CRISPR. HR-L is the homology arm upstream HXK1. HR-R is the homology arm |
| + | downstream HXK1. pgaA is the sequence of pgaA inserted in the homology arm (Figure 2). </p> |
| + | |
| + | <div class="img-container m-t-12 m-b-12"> |
| + | <img src="https://static.igem.org/mediawiki/2021/1/1b/T--Xiamen_City--img_contribution_5.jpg" alt="" style="width: 80%;"> |
| + | <span class="figure">Figure 2. Cas9+gRNA+HR-L-endo-pgaA-HR-R box.</span> |
| + | </div> |
| + | </section> |
| + | </section> |
| + | </div> |
| + | </div> |
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