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[GUIDELINES]
<p><strong>Buffers and Solutions</strong></p>
<p> </p>
<p><strong>Suspension Buffer (SB):</strong></p>
<p>25 mM Tris pH 7.5</p>
<p>1 M Sorbitol</p>
<p>25 mM EDTA pH 8.0</p>
<p> </p>
<p><strong>Digestion Buffer (DB):</strong></p>
<p>250 mM EDTA pH 9.5</p>
<p>1% N-Lauroylsarcosine<br /><br /><br /></p>
<p><strong>1 × TAE Buffer:</strong></p>
<p>40 mM Tris</p>
<p>20 mM acetic acid</p>
<p>1 mM EDTA</p>
<p> </p>
<p><strong>Formulation of MBBM</strong> (Modified Bold’s Basal Medium)</p>
<p>Stock Solutions:</p>
<ol>
<li>25.0 g NaNO<sub>3 </sub>/1L d-H<sub>2</sub>O</li>
<li>2.5 g CaCl<sub>2 </sub><strong><sup>.</sup></strong> 2H<sub>2</sub>O /1L d-H<sub>2</sub>O</li>
<li>7.5 g MgSO<sub>4</sub><strong><sup> .</sup></strong> 7H<sub>2</sub>O /1L d-H<sub>2</sub>O</li>
<li>7.5 g K<sub>2</sub>HPO<sub>4 </sub>/1L d-H<sub>2</sub>O</li>
<li>17.5 g KH<sub>2</sub>PO<sub>4 </sub>/1L d-H<sub>2</sub>O</li>
<li>2.5 g NaCl /1L d-H<sub>2</sub>O</li>
<li>50.0 g disodium EDTA; 31.0 g KOH /1L d-H<sub>2</sub>O</li>
<li>4.98 g FeSO<sub>4</sub> <strong><sup>.</sup></strong> 7H<sub>2</sub>O / 1L acidified H<sub>2</sub>O (Acidified H<sub>2</sub>O is 999.0 mL d-H<sub>2</sub>O + 1.0 mL concentrated H<sub>2</sub>SO<sub>4</sub>)</li>
<li>11.42 g H<sub>3</sub>BO<sub>3</sub> /1L d-H<sub>2</sub>O</li>
<li>8.82 g ZnSO<sub>4</sub> <strong><sup>. </sup></strong>7H<sub>2</sub>O; 1.44 g MnCl<sub>2</sub> <strong><sup>.</sup></strong> 4H<sub>2</sub>O; 0.71 g MoO<sub>3</sub>; 1.57 g CuSO<sub>4</sub> <strong><sup>.</sup></strong> 5H<sub>2</sub>O; 0.49 g CoNO<sub>3</sub> <strong><sup>.</sup></strong> 6H<sub>2</sub>O /1L d-H<sub>2</sub>O</li>
</ol>
<p> </p>
<p>MBBM preparation:</p>
<p>to 950 mL of  d-H<sub>2</sub>O add:</p>
<p>10.0 mL of stock solutions 1; 2; 3; 4; 5 and 6</p>
<p>1.0 mL of stock solutions 7; 8 and 9</p>
<p>1.0 mL of stock solution 10</p>
<p>1.0 g of bacto-peptone</p>
<p>5.0 g of sucrose</p>
<p> </p>
<p><strong>Formulation of FES Medium </strong>(for Chlorella Pbi growth)</p>
<p>Stock Solutions:</p>
<p>1). 10.0 gm MgSO<sub>4</sub><strong><sup>.</sup></strong>7H<sub>2</sub>O/1L d-H<sub>2</sub>O</p>
<p>2). 1.0 gm KNO<sub>3</sub>/1L d-H<sub>2</sub>O</p>
<p>3). 1.0 gm K<sub>2</sub>HPO<sub>4 </sub>/1L d-H<sub>2</sub>O</p>
<p>4). 50.0 g disodium EDTA; 31.0 g KOH /1L d-H<sub>2</sub>O</p>
<p>5). 4.98 g FeSO<sub>4</sub> <strong><sup>.</sup></strong> 7H<sub>2</sub>O / 1L acidified H<sub>2</sub>O (Acidified H<sub>2</sub>O is 999.0 mL  d-H<sub>2</sub>O + 1.0   </p>
<p>      mL concentrated H<sub>2</sub>SO<sub>4</sub>)</p>
<p>6). 11.42 g H<sub>3</sub>BO<sub>3</sub> /1L d-H<sub>2</sub>O</p>
<p>7). 8.82 g ZnSO<sub>4</sub> <strong><sup>. </sup></strong>7H<sub>2</sub>O; 1.44 g MnCl<sub>2</sub> <strong><sup>.</sup></strong> 4H<sub>2</sub>O; 0.71 g MoO<sub>3</sub>; 1.57 g CuSO<sub>4</sub> <strong><sup>.</sup></strong> 5H<sub>2</sub>O; </p>
<p>      0.49 g CoNO<sub>3</sub> <strong><sup>.</sup></strong> 6H<sub>2</sub>O /1L d-H<sub>2</sub>O</p>
<p> </p>
<p>FES  preparation:</p>
<p>to 950 mL of  d-H<sub>2</sub>O add:</p>
<p>20.0 mL of stock solutions 1, 2, and 3.</p>
<p>1.0 mL of stock solutions 4, 5  and 6.</p>
<p>2.0 mL of stock solution 7</p>
<p>1.0 g of bacto-peptone</p>
<p>2.0 gm of Oxoid Lab-Lemco Powder</p>
<p>5.0 g of sucrose</p>
<p> </p>

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