YLEX Expression Kit

To shorten the experimental time and simplify the procedures.

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YLEX Expression Kit (download PDF)

YLEX Expression Kit based on INRA INAPG licensed patent* provides an easy approach for cloning and expressing a gene of interest in the yeast, Yarrowia lipolytica . Using this kit, high level of heterologous protein may be expressed intracellularly or be secreted from the cell into medium by selecting the supplied expression vector pYLEX1 or pYLSC1.

Vectors provided in the YLEX contain a strong hybrid promoter carrying four tandem copies of upstream activator sequences (UAS1B) fragment from pXPR2 and a minimal pLEU2 fragment. Unlike the frequently used Yarrowia promoter (p XPR2 ), this stable hybrid promoter directs protein expression constantly without multiple influences by nutritional and environmental factors in medium.

When a constructed plasmid with the hybrid promoter followed by a cloned gene of interest is linearized by the selected restriction enzyme, it becomes an expression cassette that can integrate into the Y. lipolytica genome by homologous recombination within the process of transformation. The successful transformants are ready for expression or secretion of recombinant protein depending on whether secretion signal appears on the plasmid.

For more information, please read the articles cited in this catalog.

※INRA (Institut National de la Recherche Agronomique) and INAPG (Institut National Agronomique Paris-Grignon)

Features and Benefits

※	Safe :Yarrowia lipolytica was classified as GRAS (generally regarded as safe) by the US FDA (Food and Drug Administration)
※	Simple : A simple tool for expressing heterologous protein
※	Easy manipulation : Like E. coli and Saccharomyces cerevisiae
※	Stable : Strong stability in vectors and constructed plasmids
※	Reliable : Vectors integrated at the same site in genome
※	Flexible : Both expression and secretion vector provided (Proteins may be expressed intracellularly or be secreted from the cell into medium)
※	High growth ability, High secretion capacity & High product yield
※	Less protein degraded : No extracellular protease synthesized by a special protease-deficient Yarrowia strain
※	Mass Production : Industrial mass production of recombinant proteins
※	Less hyperglycosylation : Able to perform post-translational processing of complex proteins, unlike Saccharomyces cerevisiae

A Yeast Strain

The strain Po1g of Yarrowia lipolytica is a derivative of the wild-type strain W29 (ATCC 20460) by a series of genetic modifications.

Yarrowia Vectors

Two vectors (pYLEX1 and pYLSC1) are included in this kit, and they can be used for either intracellular expression or secretion of proteins of interest in Y. lipolytica . Generally speaking, if the target protein is cytosolic and non-glycosylated, the pYLEX1 vector is a better choice. If the protein of your interest is normally glycosylated or secreted, you may wish to choose the pYLSC1 vector.

The pYLEX1 expression vector (7259 bp) contains the strong hybrid promoter (hp4d) carrying four tandem copies of upstream activator sequences (UAS1B) fragment from p XPR2 and a minimal p LEU2 fragment. The multiple cloning site and the XPR2 transcription terminator lie immediately downstream of 3' site of hp4d promoter, followed by a leucine selection marker gene ( LEU2 ). The vector can be linearized by digestion with Not I to create a linear DNA fragment capable of inserting into the Y. lipolytica genome.

The pYLSC1 secretion vector (7205 bp) contains the hybrid promoter (hp4d) and a secretion signal ( XPR2 pre region). The multiple cloning site and the p XPR2 transcription terminator lie immediately downstream of 3' site of XPR2 pre region, followed by a leucine selection marker gene ( LEU2 ). The vector can be linearized by digestion with Not I to create a linear DNA fragment capable of inserting into the Y. lipolytica genome.

Strategic Procedure

Experimental Data

Detection of recombinant α-amylase secreted by Yarrowia transformants
The figure shows that filtered culture medium from batch culture of both amylase-encoding transformants (YL #2 and #3) could digest starch in solid medium agar, and subsequently produce clear zones. In contrast, medium from the culture of yeast transformed with vector only (YL #1) did not exhibit the same result. It indicates that cloning of both α-amylase gene ( AMY1 ) into pYLEX1 and a -amylase gene without its secretion signal peptide ( AMY1 D ) into pYLSC1 was successful by using the YLEX Expression Kit. In both cases, active α-amylase was efficiently secreted into the culture medium.

Ordering Information

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