2A

2Apeptide sequenceswere found in Picornaviruses to mediate "cleavage" between two proteins.We use 2A peptide-linked multicistronic vectors to express multiple proteins from a single open reading frame (ORF)effectively.
The 18~22 amino acids 2A self-cleaving oligopeptidescan be used for co-expression ofmultiple, discrete proteins from a single ORF.Based on highly inefficient peptide bond formation between glycineand proline residues within the 2A peptide, placementof 2A peptide sequence as a linker region betweentandem cDNA’s allows the stoichiometric translation ofmultiple unfused protein products.These sequences were first discovered in the foot-and-mouth disease virus (FMDV).And since than many 2A-like sequences have been identified in other viruses and some parasites.To minimize therisk of homologous recombination, it is important to use different 2A peptide sequences if morethan two genes are being linked.The 2A peptide system has thus far worked successfully in all eukaryotic systems tested, from mammaliancells, yeast, and plants.In our project,we use F2A(from foot-and-mouth disease virus), P2A(from porcine teschovirus-1) and T2A(fromThosea asigna virus) to achieve our goal.

GSG linker

GSG linker is anoligopeptide of “Gly-Ser-Gly” between your protein and 2A peptide to enhance cleavage.
Ribosomal “skipping” is an alternate mechanism of translation in which a specific viral peptide prevents the ribosome from covalently linking a new inserted aa, and let it continue translation. This result in apparent co-translational cleavage of the polyprotein.


This process is induced by a “2A-like”, or CHYSEL (cis-acting hydrolase element) sequence. This sequence comprises a non-conserved sequence of amino-acids with a strong alpha-helical propensity followed by the consensus sequence -D(V/I)ExNPG P, where x= any amino acid. The apparent cleavage occurs between G and P.
Ribosomal “skipping” has been observed only in +ssRNA and dsRNA viruses, whose hosts are animals, plant or insect. The conserved motif has been identified also in trypanosoma and in some fungus proteins. Ribosomal “skipping” is functioning both in vitro and in vivo when translated by eukaroytic ribosomes, but is inactive when translated by prokaryoticribosomes.

GSG linker