Synthetic expression vectors for insect cells
View Patent ↗The present invention is directed at optimized expression vectors for the expression of native-like heterologous proteins in insect cells. Compositions of the invention are nucleotide sequences representing elements of an expression vector that when combined results in enhanced expression and secretion of heterologous proteins. The elements include sequences that define transcriptional activators, core promoters, secretion signals, and 3′ untranslated regions that are functional in insect cells. The elements contained in the optimized vectors are all synthetically derived or are modified variants of naturally occurring insect sequences. The expression vectors are useful for the expression of native-like proteins when protein encoding nucleotide sequences are operatively linked to the vectors. These vectors can be used to transform insect cells, which can then be cultured to produce the desired protein product. The expressed native-like proteins can be used in diagnostic, vaccine or other applications requiring large amounts of high quality proteins.
1. An expression vector for expression and secretion of heterologous proteins in cultured insect cells, comprising an expression cassette, wherein the expression cassette contains a synthetic promoter that is composed of proximal and core promoter elements that are operably linked within the expression cassette such that the insertion of a heterologous gene sequence will result in protein expression from cells stably transformed with the expression vector wherein the proximal promoter comprises the 3XR MRE-3XR MRE sequence shown in SEQ ID NO:6 and the core promoter comprises the SCP1-TolloMTEΔ111 sequence shown in SEQ ID NO: 2 or the SCP7 sequence shown in SEQ ID NO: 3.
2. The expression vector of claim 1 , further comprising a synthetic 5′UTR element operably linked to the synthetic proximal promoter and core promoter such that the combination is capable of driving the expression of heterologous proteins.
3. The expression vector of claim 2 , further comprising a synthetic secretion signal sequence operably linked to the synthetic proximal promoter, core promoter and 5′UTR elements such that the combination is capable of driving the expression and secretion of heterologous proteins.
4. The expression vector of claim 3 , further comprising a synthetic 3′UTR sequence operably linked to the synthetic proximal promoter, core promoter, 5′UTR, and synthetic secretion signal sequence elements such that the combination is capable of driving the expression and secretion of heterologous proteins.
5. The expression vector of claim 2 , wherein the 5′UTR element comprises the sequence shown in SEQ ID NO:4.
6. The expression vector of claim 2 , wherein the 5′UTR element comprises the sequence shown in SEQ ID NO:5.
7. The expression vector of claim 3 , wherein the synthetic secretion signal sequence comprises the 1R5L(H) encoding sequence shown in SEQ ID NO:7.
8. The expression vector of claim 4 , wherein the synthetic 3′UTR comprises the Op CLP 3′UTR sequence shown in SEQ ID NO:9.
9. An expression vector for expression and secretion of heterologous proteins in cultured insect cells, comprising the expression cassette shown in SEQ ID NO:10.
10. An expression vector for expression and secretion of heterologous proteins in cultured insect cells, comprising the expression cassette shown in SEQ ID NO:11.