T7 polymerase often inserts a terminal A nucleotide to the 3' CCC sequence, however, this did not appear to significantly influence RNA template recognition.
A similar experiment with two primer pairs VRARDHIS and VRARIHIS was set up to remove the N-terminal TEV protease cleavable His-tag and insert a C-terminal His-tag in a single reaction.
Since fluorescently tagged γ-tubulin fusions were not fully functional (not shown), we used a CRISPR/Cas9-mediated strategy (Dickinson et al., 2013) to insert a C-terminal GFP tag in the endogenous locus of gip-2, which encodes an essential component of the C. elegans γ-tubulin complex (Hannak et al., 2002).
This receptor was engineered by inserting a C-terminal rho1D4 epitope tag and an N-terminal FLAG epitope tag to allow its purification and detection.
ER-GFP plasmid was made from pEGFP-C2, by inserting a C-terminal KDEL sequence by SDM, and an N-terminal ER-signal sequence from calreticulin, by oligo-annealing between NheI and AgeI restriction sites.
However, ligation of the new insert into the vector is a terminal product, because the new insert contains no BsaI sites and therefore cannot be cut by BsaI.
To create a control strain, we inserted a C-terminal 3ha tag and a TRP1 selection marker at the RPN4 locus in the parental YFP17 strain.
To target Akt1 to the membrane for constitutive activation, we inserted a C-terminal sequence tag encoding a farnesylation motif and a stretch of basic amino acids (Schmidt et al, 2002).
One simply rolls the bar-code reader across the page of a photo sample book and then inserts the device into a terminal on the camera.
You'll be inserting the card in a terminal instead of swiping it.
After 48 h of culture, transient gene expression was determined by western blotting, because the inserted sequences were ended with a terminal codon before the EGFP gene.
