... Since researchers can generate almost any DNA sequence in the way that they like, the design and engineering...mNeonGreen, mScarlet(-I), and mCherry. Keep the linker sequence short In the early days of GFP applications, many...site and therefore consist of random amino acid sequences or (ii) designed to form an inert, unstructured...and protein of interest (POI). To indicate the sequence, people often use terminology like “C-terminal...interest usually outweigh potential negative consequences of the tag. Many thanks to our guest blogger...

...N-terminal consensus sequence (MG-) and prenylation requires a C-terminal consensus sequence (-CaaX). Both modifications...modifications are disrupted when GFP is fused to these sequences. When N- and C-terminal fusions are not tolerated...alternative is to insert the GFP into the coding sequence of the protein of interest. One of the reasons... reaction requires molecular oxygen and as a consequence GFP remains non-fluorescent under anaerobic conditions...

...segments, each with two fluorescent protein coding sequences in opposite orientations. Cre-mediated excision...the system, the fluorescent protein whose coding sequence is closest to the promoter will be expressed. ...used are fluorescently distinct, their protein sequences are highly homologous, preventing design of antibodies...three with low levels of both fluorescence and sequence overlap (coral mOrange2, jellyfish EGFP, and sea...

...MG1655    Wild type K-12 strain; first published sequence of an E. coli K-12 strain. F- λ- ilvG- rfb-50 ...and retroviral vectors or cloning or repeated sequences with the potential to recombine.  F' proA+B+ lacIq...and retroviral vectors or cloning or repeated sequences with the potential to recombine.  F- endA1 glnV44...and retroviral vectors or cloning or repeated sequences with the potential to recombine. This strain is...E.coli strains Gene(s) Description Functional Consequence dam DNA adenine methylase mutation (GATC) Preparing...

...strange breaks at target sites 5' of the 5'NGG3' PAM sequence as directed by a gRNA. 2A-Puro In this plasmid...into which you can clone your 20 nucleotide guide sequence of choice (for generation of a gRNA that will ...

...phosphorylatable residues and the bNLS and NES sequences, which have previously been characterized. See...more details on the design of the bNLS and NES sequences. Disadvantages of KTRs KTRs also have some disadvantages...

...In the O'Connor lab, we recognized that genome sequencing projects were generating huge amounts of data...available. Then, staff or students clone similar sequences from a less-well studied organism into overexpression...experiments We used TOPO cloning to insert the coding sequences for ten different MET/Met genes (Figure 1) from...

...that recognize, cut, and alter specific target sequences in DNA resulting in a variety of outcomes including... requirement. For instance, for Casɸ-2 the PAM sequence is 5’-TBN-3’ (where B is G, T, or C). The authors...scientists seeking a wider range of targetable genomic sequences could benefit from CRISPR-Casɸ. Pausch et al....

...approaches: single-cell RNA-sequencing to identify neuronal class and deep sequencing of neurons from each class...of a GAL-4 driver that can bind to activation sequences upstream of genes to stimulate expression (Wang...

...strategies: Strategy #1 (ProA): Create promoters with sequences upstream of the start codon of retinal cell-type...Create promoters using phylogenetically conserved sequences that are upstream of the transcription initiation... factor binding sites interspaced with random sequences.   Strategy #4 (ProD): Create promoters with the...