pLD‐CoHSA‐Trxf

Protocol tips

Upstream tips	Protocol tips	Downstream tips
	The resulting plasmids, termed pET Trxm and pET Trxf, were introduced into E. coli (BLR DE3, Novagene).	Overexpressed proteins were purified by chromatography in affinity columns packed with Ni‐NTA agarose (Qiagen, Hilden, Germany). Specific anti‐Trx m and anti‐Trx f antibodies were raised by immunizing rabbits with purified His‐tagged proteins (Abyntek, Bilbao, Spain). Antibody cross‐reactivity was eliminated by affinity purification (exclusion) of anti‐Trx m IgG with the Trx f antigen and vice versa.

Protocol tips
The resulting plasmids, termed pET Trxm and pET Trxf, were introduced into E. coli (BLR DE3, Novagene).
Downstream tips
Overexpressed proteins were purified by chromatography in affinity columns packed with Ni‐NTA agarose (Qiagen, Hilden, Germany). Specific anti‐Trx m and anti‐Trx f antibodies were raised by immunizing rabbits with purified His‐tagged proteins (Abyntek, Bilbao, Spain). Antibody cross‐reactivity was eliminated by affinity purification (exclusion) of anti‐Trx m IgG with the Trx f antigen and vice versa.

Publication protocol

Tobacco Trx m and Trx f were expressed in E. coli as His‐tagged polypeptides. The coding sequences were amplified by PCR with the same primers as those used for their cloning (FusTrxm/f‐For and CoTrxm/f‐Rev); the resulting PCR fragment was digested with NcoI and NotI and cloned in a pET 28a(+) vector (Novagene; Merck KGaA, Darmstadt, Germany). The resulting plasmids, termed pET Trxm and pET Trxf, were introduced into E. coli (BLR DE3, Novagene). Overexpressed proteins were purified by chromatography in affinity columns packed with Ni‐NTA agarose (Qiagen, Hilden, Germany). Specific anti‐Trx m and anti‐Trx f antibodies were raised by immunizing rabbits with purified His‐tagged proteins (Abyntek, Bilbao, Spain). Antibody cross‐reactivity was eliminated by affinity purification (exclusion) of anti‐Trx m IgG with the Trx f antigen and vice versa.

Full paper   Login or join for free to view the full paper.

Reviews

Discussion

Papers

Check out relevant papers found by Labettor's AI that are relevant for performing Protein Expression Prokaryotic cells - E. coli Tobacco Trxf using pLD‐CoHSA‐Trxf from Inmaculada Farran, Instituto de Agrobiotecnología (UPNA‐CSIC�.

Manufacturer protocol

Download the product protocol from Inmaculada Farran, Instituto de Agrobiotecnología (UPNA‐CSIC� for pLD‐CoHSA‐Trxf below.

Videos

Check out videos that might be relevant for performing Protein Expression Prokaryotic cells - E. coli Tobacco Trxf using pLD‐CoHSA‐Trxf from Inmaculada Farran, Instituto de Agrobiotecnología (UPNA‐CSIC�. Please note that these videos are representative and steps or experiment specific processes must be kept in mind to expect desired results.