Team:Chiba/Project

From 2013.igem.org

(Difference between revisions)

Revision as of 17:24, 27 September 2013

iGEM-2013 Chiba

Introduction

Why bother magnetizing?
We are surrounded by magnets. Magnetized materials are everywhere makes up core components of electrical switches, speaker systems, and portable memory devices (including credit cards), and diagnostic/ separation systems. Our hope is to program E. coli cell to turn into magnets to be used for various purposes.

Magnet force is unique in that;
1. Non-contact (least chance of damaging and contamination)
2. Barrier-free mode of collections (can penetrate the physical blockage)
3. Rapidity (immediate exercise)
4. Duration (eternal action if needed)

What is in future?
We decided to design & implement biobricks that are effective for magnetizing E. coli.
If we can freely magnetize any given cells, together with the cell-surface display techniques, you can establish novel systems for bioseparations. Or, you can simplify the harvesting/ collecting step in the process of bioproduction and bioremediation.

Overview/Strategies

    We have three steps to create magnetic E. coli.

1. Reprogramming the iron homeostasis:To maximize the chance of magnetization, we would like to pump as much Fe into the cell as possible, and keep it. To this end, we tried to eliminate the negative controller (encoded by fur) on the Fec system (iron importer). Also, we tried to knock down the Fe exporter fieF.

    Some metal oxides of the spinel type i.e. Fe₃O₄ have ferrimagnetism because of the disparity of magnetic moment. If E. coli has it, it will be attracted by magnets.
    Ordinary, Fe inside cytosol are reduced and exists as divalent ferrous ion. Fe ions, which injure the DNAs with hydroxyl radical from Fenton reaction, are isolated from cytosol by ferritin. Ferritin accumulates ferrous ions inside and oxidizes to ferric ions for prevention of Fenton reaction.
    Fur controls E. coli iron metabolism. In the case of iron oversupply, fur combine with ferrous ion and restricts the expression of Fec(iron importer) and induces the expression of ferritin. In the other hand, fieF is iron efflux pump which fur doesn't control. These maintain iron homeostasis of E. coli.
    In our project, (1) we knock out genes, trxB and gor to shift the oxidation state to more oxidized state inside cytosol so that ferrous ions are easier to be oxidized and exist in cytosol. And, (2) human ferritins are overexpressed in order to deal with ferrous ions quickly which haven't be oxidized yet. It will improve iron tolerance of E. coli. (3)We knock down fur and fieF by CRISPRi for the sake of increase the amount of iron uptake.

We have three steps to create magnetic E. coli.

increase iron uptake

improve iron storage

change the iron redox state

And, there are each analytical methods.

Project

Iron Uptake	Iron Storage	Iron Oxidation

@@ Line 38: / Line 38: @@
 <div class="section">
-	<h2 id="over" style="background-color:#ff9933"><center>Overview</center></h2>
+	<h2 id="over" style="background-color:#ff9933"><center>Overview/Strategies</center></h2>
-	<p>&nbsp;&nbsp;&nbsp;&nbsp;Some metal oxides of the spinel type <i>i.e.</i> Fe<sub>3</sub>O<sub>4</sub> have ferrimagnetism because of the disparity of magnetic moment. If <i>E. coli</i> has it, it will be attracted by magnets.
+	<p>&nbsp;&nbsp;&nbsp;&nbsp;We have three steps to create magnetic E. coli.<br><br>
+<strong>1. Reprogramming the iron homeostasis:</strong>To maximize the chance of magnetization, we would like to pump as much Fe into the cell as possible, and keep it.  To this end, we tried to eliminate the negative controller (encoded by fur) on the Fec system (iron importer). Also, we tried to knock down the Fe exporter fieF. <br><br>
+&nbsp;&nbsp;&nbsp;&nbsp;Some metal oxides of the spinel type <i>i.e.</i> Fe<sub>3</sub>O<sub>4</sub> have ferrimagnetism because of the disparity of magnetic moment. If <i>E. coli</i> has it, it will be attracted by magnets.
 	<br>&nbsp;&nbsp;&nbsp;&nbsp;Ordinary, Fe inside cytosol are reduced and exists as divalent ferrous ion. Fe ions, which injure the <i>DNA</i>s with hydroxyl radical from Fenton reaction, are isolated from cytosol by ferritin. Ferritin accumulates ferrous ions inside and oxidizes to ferric ions for prevention of Fenton reaction.
 	<br>&nbsp;&nbsp;&nbsp;&nbsp;<i>Fur</i> controls <i>E. coli</i> iron metabolism. In the case of iron oversupply, fur combine with ferrous ion and restricts the expression of <i>Fec</i>(iron importer) and induces the expression of ferritin. In the other hand, <i>fieF</i> is iron efflux pump which fur doesn't control. These maintain iron homeostasis of <i>E. coli</i>.