The growth complementation assays were performed as previously described74 (link). In brief, E. coli LB2003, a strain lacking all endogenous K+ uptake systems, was transformed with plasmids encoding His-tagged KdpFABC variants. LB2003 transformed with empty vector pBAD18 or plasmid pBXC3H-KdpFABD307NC, encoding for an inactive variant, served as negative controls. Growth was monitored for 24 h at different K+ concentrations (1–115 mM, referred to as K1–K115). At K10 and below, the strain only grows sufficiently if the produced protein complements the lacking K+ transport systems. Protein production was confirmed by SDS-PAGE and subsequent western blotting analysis of a K30 sample after 24 h using an anti-His antibody from mouse (dilution 1:3000, Sigma–Aldrich, cat.no. H1029) and secondary anti-mouse IgG-HRP antibody produced in goat (dilution 1:20,000, Sigma–Aldrich, cat.no. A2554).
Anti his antibody from mouse
Manufactured by Merck Group
The Anti-His antibody from mouse is a laboratory reagent used for the detection and purification of recombinant proteins containing a histidine (His) tag. It is a highly specific antibody that binds to the His tag, allowing the identification and isolation of the tagged protein from complex samples.
Automatically generated - may contain errors
Lab products found in correlation
3 protocols using anti his antibody from mouse
1
K+ Uptake System Complementation Assay
2
Complementation Assay for K+ Transport Systems
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The growth complementation assays were performed as previously described29 (link). In brief, His-tagged versions of KdpA, KdpFABC and KtrB, respectively, were expressed in E. coli LB2003, a strain lacking all endogenous K+ uptake systems. LB2003 transformed with empty vector pBAD18 served as negative control. Growth curves were recorded for 24 h at different K+ concentrations (1–115 mM referred to as K1–K115). At K10 and below the strain only grows sufficiently, if the expressed protein complements the lacking transport systems. Protein production was confirmed by Western blotting analysis of a K30 sample after 24 h using an anti-His antibody from mouse (dilution 1:3000, Sigma-Aldrich, cat.no. H1029) and secondary anti-mouse IgG-peroxidase antibody produced in goat (dilution 1:20,000, Sigma-Aldrich, cat.no. A2554).
3
Complementation Assay for K+ Transport
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The growth complementation assays were performed as previously described 66 .
In brief, E. coli LB2003, a strain lacking all endogenous K + uptake systems, was transformed with plasmids encoding His-tagged KdpFABC variants. LB2003 transformed with empty vector pBAD18 or plasmid pBXC3H-KdpFABD307NC, encoding for an inactive variant, served as negative controls. Growth was monitored for 24 h at different K + concentrations (1-115 mM, referred to as K1-K115). At K10 and below, the strain only grows sufficiently if the produced protein complements the lacking K + transport systems. Protein production was confirmed by SDS-PAGE and subsequent Western blotting analysis of a K30 sample after 24 h using an anti-His antibody from mouse (dilution 1:3000, Sigma-Aldrich, cat.no.
H1029) and secondary anti-mouse IgG-HRP antibody produced in goat (dilution 1:20,000, Sigma-Aldrich, cat.no. A2554).
In brief, E. coli LB2003, a strain lacking all endogenous K + uptake systems, was transformed with plasmids encoding His-tagged KdpFABC variants. LB2003 transformed with empty vector pBAD18 or plasmid pBXC3H-KdpFABD307NC, encoding for an inactive variant, served as negative controls. Growth was monitored for 24 h at different K + concentrations (1-115 mM, referred to as K1-K115). At K10 and below, the strain only grows sufficiently if the produced protein complements the lacking K + transport systems. Protein production was confirmed by SDS-PAGE and subsequent Western blotting analysis of a K30 sample after 24 h using an anti-His antibody from mouse (dilution 1:3000, Sigma-Aldrich, cat.no.
H1029) and secondary anti-mouse IgG-HRP antibody produced in goat (dilution 1:20,000, Sigma-Aldrich, cat.no. A2554).
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!