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--- purification:expression_and_purification_of_smprmt3_184-553 [2016/10/28 13:36] – marechan
+++ purification:expression_and_purification_of_smprmt3_184-553 [2023/01/20 13:33] (current) – removed marechan
@@ Line 1: / Line 1: @@
-**__Expression & purification of 6his-SmPRMT3(1-553)__**
-**Construct**
-MGSSHHHHHHSSGTGSGENLYFQGHMSPCDQSSNYDLDMEPNDSSYFGSYGHFEIHGEMINDRVRTESYVNFILSNAEKYFKHKIILDVGSGSGILSIIAAQAGASHVYGVEAADEIYAASHETLRVNNLLERVTFIHGQAESVELPVKKVDVIISEWMGYFLFFESMLDSVLKMASKYLSRDGHIFPRHYTLNLLGVQCSEQLRKRRLEHWNNVYGYNMPALRRAALSEAHVLNLTNEHVTPPISPITILTQSFELVALDLDDMHRNRIYNLSNHCSLLCEQKFHLTIQPTTDINNNSSSSSYELDAIVGYFDVRFDDDADCKVEFSTSPTTPLTHWKQTLLFLDKPIRVKPGDKISGIITIRRATTDNRGLEINLLIGETENSLEIKQTFDLIG
-Length = 396 AA
-Molecular weight = 44782.3 Da
-pI 5.66
-ξ(red) 40340 L.M<sup>-1</sup>
-**Expresion**
-Grow pnEAvH_SmPRMT3-184 BL21(DE3) transformants on an LB-Amp plate (1 plate/L culture), overnight 37°C. Resuspend cells in LB medium (5 mL/plate) and innoculate liquide **2X** LB medium + 100 μg/mL<sup>-1</sup> Ampicilline. Grow culture(s) at 37°C, 200 rpm and measure OD600 every 20 minutes. When OD600 reaches 0.4, set temperature to 20°C. When OD600 reaches 0.6 to 0.8, induce 6his-SmPRMT3(184-553) with 1 mM IPTG and leave the cultures to express the protein overnight.