CPMV enhancer elements
An expression enhancer comprising a CPMV 5′UTR nucleotide sequence consisting of X nucleotides (CMPVX), where X=160, 155, 150, or 114 of SEQ ID NO:1, or consisting of a nucleotide sequence comprising from about 80% to 100% sequence similarity with CMPVX, where X=160, 155, 150, or 114 of SEQ ID NO:1 SEQ ID NO:1 is provided. The expression enhancer may further comprise a stuffer sequence fused to the 3′ end of the 5′UTR nucleotide sequence (CMPVX+, where X=160, 155, 150, or 114 of SEQ ID NO:1). The stuffer sequence may comprise one or more plant kozak sequences. Plants comprising the expression enhancer and methods using the expression enhancer are also described.
1. A nucleic acid molecule comprising:
a first portion consisting of nucleotides 1-160 of SEQ ID NO:1 having a substitution of G1 15A, and
a second portion encoding a heterologous protein of interest, the second portion operatively linked to and fused immediately to the 3′ end of the first portion,
wherein transient expression of the nucleic acid molecule in a Nicotiana benthamiana plant increases a yield of the heterologous protein of interest, compared to transient expression of a comparative nucleic acid molecule comprising a corresponding first portion consisting of SEQ ID NO:4 in a comparative plant.
2. A plant expression system comprising the nucleic acid molecule of claim 1 or a vector comprising the nucleic acid molecule of claim 1 .
3. The plant expression system of claim 2 , wherein the nucleic acid molecule or vector further comprises a comovirus 3′ UTR.
4. The plant expression system of claim 2 , wherein the heterologous protein of interest is a viral protein or an antibody.
5. The plant expression system of claim 4 , wherein the viral protein is an influenza hemagglutinin selected from the group consisting of H1, H2, H3, H4, H5, H6, H7, H8, H9, H10, H11, H12, H13, H14, H15, H16, and influenza type B hemagglutinin.
6. A method of producing a protein of interest in a plant or in a portion of a plant comprising, introducing into the plant or in the portion of a plant the plant expression system of claim 2 , and incubating the plant or the portion of a plant under conditions that permit expression of the heterologous protein of interest.
7. A plant or portion of a plant transiently transfected or stably transformed with the plant expression system of claim 2 .
8. The nucleic acid molecule of claim 1 , wherein the heterologous protein of interest is a viral protein or an antibody.
9. The nucleic acid molecule of claim 8 , wherein the viral protein is an influenza hemagglutinin selected from the group consisting of H1, H2, H3, H4, H5, H6, H7, H8, H9, H10, H11, H12, H13, H14, H15, H16, and influenza type B hemagglutinin.
10. The plant expression system of claim 2 , further comprising a second nucleic acid molecule, the second nucleic acid molecule comprising a nucleotide sequence encoding a suppressor of silencing.
11. The plant expression system of claim 10 wherein the suppressor of silencing is selected from the group HcPro and p19 suppressor of silencing.