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Team:Evry/Biology/CellCharacterization/Culture
From 2014.igem.org
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<h2>Cell culture</h2> | <h2>Cell culture</h2> | ||
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With the help of the 'Molecules of defence and communication in the microbial ecosystems' team of the National museum of natural history of Paris (their web page <a href="http://mcam.mnhn.fr/MDCEM/index.htm">here</a>), we succeed to cultivate our bacteria <i>Pseudovibrio denitrificans</i> in two medium, the marine broth (MB) and the minimal medium M9 added 2% of CASAmino Acid and by 3% of NaCl.<br> | With the help of the 'Molecules of defence and communication in the microbial ecosystems' team of the National museum of natural history of Paris (their web page <a href="http://mcam.mnhn.fr/MDCEM/index.htm">here</a>), we succeed to cultivate our bacteria <i>Pseudovibrio denitrificans</i> in two medium, the marine broth (MB) and the minimal medium M9 added 2% of CASAmino Acid and by 3% of NaCl.<br> | ||
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| - | <b><u>Table1:</u></b> Composition of used media, Marine Broth and M9-CASA+3% NaCl.<br> | + | <p><b><u>Table1:</u></b> Composition of used media, Marine Broth and M9-CASA+3% NaCl.<br></p> |
<img src="https://static.igem.org/mediawiki/2014/a/ac/Table_of_media4.jpg" alt="text to print if image not found" /><br> | <img src="https://static.igem.org/mediawiki/2014/a/ac/Table_of_media4.jpg" alt="text to print if image not found" /><br> | ||
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Because of the opaqueness of the marine broth, it's impossible to obtain reliable data of the growth of the bacteria. Thus, the growth curve that we have made, has been made in M9-CASA+3%NaCl 1X, with the <a id="igem-link" target="_blank" href="http://www.tecan.com/platform/apps/product/index.asp?MenuID=1813&ID=1918&Menu=1&Item=21.2.10.1.1 " title="Go to iGEM site">TECAN infiniteM200</a> (cf: Figure1). <br> | Because of the opaqueness of the marine broth, it's impossible to obtain reliable data of the growth of the bacteria. Thus, the growth curve that we have made, has been made in M9-CASA+3%NaCl 1X, with the <a id="igem-link" target="_blank" href="http://www.tecan.com/platform/apps/product/index.asp?MenuID=1813&ID=1918&Menu=1&Item=21.2.10.1.1 " title="Go to iGEM site">TECAN infiniteM200</a> (cf: Figure1). <br> | ||
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<img src="https://static.igem.org/mediawiki/2014/2/2c/Microscope.png" alt="Curve TECAN+photo microscope?" /><br> | <img src="https://static.igem.org/mediawiki/2014/2/2c/Microscope.png" alt="Curve TECAN+photo microscope?" /><br> | ||
| - | <b><u>Figure1:</u></b> Curve of growth in liquid M9-CASA+3%NaCl 1X and microscopic photo of <i>Pseudovibrio denitrificans</i> (left) and <i>Pseudovibrio ascidiaceicola</i> (right).<br> | + | <p><b><u>Figure1:</u></b> Curve of growth in liquid M9-CASA+3%NaCl 1X and microscopic photo of <i>Pseudovibrio denitrificans</i> (left) and <i>Pseudovibrio ascidiaceicola</i> (right).<br></p> |
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The marine broth is more complete than the M9-CASA+3%NaCl, even if we have adjusted the salinity. As we can see in the Table2, <i>Pseudovibrio denitrificans</i> grows better on MB media. The number of colony is higher, and they are significantly bigger (cf: Figure2). Moreover, the growth is faster. <br> | The marine broth is more complete than the M9-CASA+3%NaCl, even if we have adjusted the salinity. As we can see in the Table2, <i>Pseudovibrio denitrificans</i> grows better on MB media. The number of colony is higher, and they are significantly bigger (cf: Figure2). Moreover, the growth is faster. <br> | ||
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<img src="https://static.igem.org/mediawiki/2014/4/4b/TableCFU.jpg" alt="Table+Photos of MB 1X and M9-CASA+3%NaCl 1X plates" /><br> | <img src="https://static.igem.org/mediawiki/2014/4/4b/TableCFU.jpg" alt="Table+Photos of MB 1X and M9-CASA+3%NaCl 1X plates" /><br> | ||
| - | <b><u>Figure2:</u></b> Number of CFU of MB 1X and M9-CASA+3%NaCl 1X plates.<br> | + | <p><b><u>Figure2:</u></b> Number of CFU of MB 1X and M9-CASA+3%NaCl 1X plates.<br> |
| - | The number of CFU was calculated by inoculated respectively 10µL of pre-cultures of <i>Pseudovibrio denitrificans</i> in MB 1X and in M9-CASA+3%NaCl at OD (600nm) ~ 1.<br> | + | The number of CFU was calculated by inoculated respectively 10µL of pre-cultures of <i>Pseudovibrio denitrificans</i> in MB 1X and in M9-CASA+3%NaCl at OD (600nm) ~ 1.<br></p> |
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| - | In the aim to know if we can transfer <i>Pseudovibrio denitrificans</i> from a media, to the other one, we have made some tests and calculate a ratio of survivability. We have made two pre-cultures of <i>Pseudovibrio denitrificans</i> in MB 1X and M9-CASA+3%NaCl 1X, and after a culture overnight, we sowed them on plates of MB 1X and M9-CASA+3%NaCl 1X. The CFU on each plate were counted and ratio of survivability were calculated using the formula Sample/Control (controls being the number of CFU obtained on a plate from a pre-culture in the same medium). (cf: Figure3). <br> | + | In the aim to know if we can transfer <i>Pseudovibrio denitrificans</i> from a media, to the other one, we have made some tests and calculate a ratio of survivability. We have made two pre-cultures of <i>Pseudovibrio denitrificans</i> in MB 1X and M9-CASA+3%NaCl 1X, and after a culture overnight, we sowed them on plates of MB 1X and M9-CASA+3%NaCl 1X. The CFU on each plate were counted and ratio of survivability were calculated using the formula Sample/Control (controls being the number of CFU obtained on a plate from a pre-culture in the same medium). (cf: Figure3). <br></p> |
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<img src="https://static.igem.org/mediawiki/2014/6/6b/Tablesiurvivability2.jpg" alt="Table+Photos of ratio of survivability" /><br> | <img src="https://static.igem.org/mediawiki/2014/6/6b/Tablesiurvivability2.jpg" alt="Table+Photos of ratio of survivability" /><br> | ||
| - | <b><u>Figure3:</u></b> Tests of survivability, table of ratio and photos.<br> | + | <p><b><u>Figure3:</u></b> Tests of survivability, table of ratio and photos.<br></p> |
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<i>Pseudovibrio denitrificans</i> can survive even if we tranfer them from a media to the other one. But we can see that even cells come from a pre-culture of M9-CASA+3%NaCl, they grow better and faster on MB plates. Moreover, even if they come from a MB pre-culture, they don't grow very well on M9-CASA+3%NaCl plates.<br> | <i>Pseudovibrio denitrificans</i> can survive even if we tranfer them from a media to the other one. But we can see that even cells come from a pre-culture of M9-CASA+3%NaCl, they grow better and faster on MB plates. Moreover, even if they come from a MB pre-culture, they don't grow very well on M9-CASA+3%NaCl plates.<br> | ||
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Revision as of 22:22, 16 October 2014
Cell culture
With the help of the 'Molecules of defence and communication in the microbial ecosystems' team of the National museum of natural history of Paris (their web page here), we succeed to cultivate our bacteria Pseudovibrio denitrificans in two medium, the marine broth (MB) and the minimal medium M9 added 2% of CASAmino Acid and by 3% of NaCl.
Table1: Composition of used media, Marine Broth and M9-CASA+3% NaCl.
Because of the opaqueness of the marine broth, it's impossible to obtain reliable data of the growth of the bacteria. Thus, the growth curve that we have made, has been made in M9-CASA+3%NaCl 1X, with the TECAN infiniteM200 (cf: Figure1).
Figure1: Curve of growth in liquid M9-CASA+3%NaCl 1X and microscopic photo of Pseudovibrio denitrificans (left) and Pseudovibrio ascidiaceicola (right).
The marine broth is more complete than the M9-CASA+3%NaCl, even if we have adjusted the salinity. As we can see in the Table2, Pseudovibrio denitrificans grows better on MB media. The number of colony is higher, and they are significantly bigger (cf: Figure2). Moreover, the growth is faster.
Figure2: Number of CFU of MB 1X and M9-CASA+3%NaCl 1X plates.
The number of CFU was calculated by inoculated respectively 10µL of pre-cultures of Pseudovibrio denitrificans in MB 1X and in M9-CASA+3%NaCl at OD (600nm) ~ 1.
In the aim to know if we can transfer Pseudovibrio denitrificans from a media, to the other one, we have made some tests and calculate a ratio of survivability. We have made two pre-cultures of Pseudovibrio denitrificans in MB 1X and M9-CASA+3%NaCl 1X, and after a culture overnight, we sowed them on plates of MB 1X and M9-CASA+3%NaCl 1X. The CFU on each plate were counted and ratio of survivability were calculated using the formula Sample/Control (controls being the number of CFU obtained on a plate from a pre-culture in the same medium). (cf: Figure3).
Figure3: Tests of survivability, table of ratio and photos.
Pseudovibrio denitrificans can survive even if we tranfer them from a media to the other one. But we can see that even cells come from a pre-culture of M9-CASA+3%NaCl, they grow better and faster on MB plates. Moreover, even if they come from a MB pre-culture, they don't grow very well on M9-CASA+3%NaCl plates.
