Difference between revisions of "Part:BBa K3580200:Experience"

 
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===Applications of BBa_K3580200===
 
===Applications of BBa_K3580200===
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Expression and purification of alanine racemase
  
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In our project, we inserted this part between bacteriophage T5 promoter for <i>E. coli</i> RNA polymerase and lambda t0 terminator of pCA24N(-gfp) planmid. This plasmid was transformed into the BL21(DE3) star strain. As this vector is chloramphenicol resistant, we always cultured the bacteria in media containing chloramphenicol. After expression induction by IPTG, the BL21 cells were collected by centrifugation, then sonicated and His-Tag purification was performed. (wikiマテメソハイパーリンク) The His-Tag purified Alr was confirmed with SDS-PAGE(Fig.1). The meanings of each abbreviation are given in Table 1. The concentration of purified racemase was quantified with the Bradford method.
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[[File:T--Waseda--racemase_parts_purification_sds.png|500px|thumb|center|Fig.1 SDS-PAGEs of purified <i>E. coli</i> BL21(DE3)star containing BBa_K3580200]]
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[[File:T--Waseda--racemase_parts_table1.png|400px|thumb|center|Table.1 Each meaning of abbriviation ]]
 
===User Reviews===
 
===User Reviews===
 
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Revision as of 07:54, 27 October 2020


This experience page is provided so that any user may enter their experience using this part.
Please enter how you used this part and how it worked out.

Applications of BBa_K3580200

Expression and purification of alanine racemase

In our project, we inserted this part between bacteriophage T5 promoter for E. coli RNA polymerase and lambda t0 terminator of pCA24N(-gfp) planmid. This plasmid was transformed into the BL21(DE3) star strain. As this vector is chloramphenicol resistant, we always cultured the bacteria in media containing chloramphenicol. After expression induction by IPTG, the BL21 cells were collected by centrifugation, then sonicated and His-Tag purification was performed. (wikiマテメソハイパーリンク) The His-Tag purified Alr was confirmed with SDS-PAGE(Fig.1). The meanings of each abbreviation are given in Table 1. The concentration of purified racemase was quantified with the Bradford method.

Fig.1 SDS-PAGEs of purified E. coli BL21(DE3)star containing BBa_K3580200
Table.1 Each meaning of abbriviation

User Reviews

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