Vectors and methods for high throughput co-expression

1. A polynucleotide comprising a first nucleotide sequence operably encoding zeomycin resistance and a second nucleotide sequence operably encoding a ccdB polypeptide, wherein the first nucleotide sequence and the second nucleotide sequence are flanked within the same attR1 and attR2 sites, and wherein the polynucleotide comprises a G144704 cassette having SEQ ID NO:4.

2. The polynucleotide of claim 1 , wherein the polynucleotide is selected from the group consisting of pDEST-C1 (SEQ ID NO:1), pDEST-C2 (SEQ ID NO:2), pDEST-C3 (SEQ ID NO:3), pDEST-CS (SEQ ID NO:17), pDEST-CS1 (SEQ ID NO:18), pDEST-CS2 (SEQ ID NO:19), pDEST-CS3 (SEQ ID NO:20), pDEST-CS4 (SEQ ID NO:21), pDEST-CMZ1 (SEQ ID NO:22), and pDEST-CMZc1 (SEQ ID NO:23).

3. An expression vector comprising the polynucleotide of claim 1 .

4. A host cell comprising the expression vector of claim 3 .

5. A polynucleotide comprising a first nucleotide sequence operably encoding tetracycline resistance and a second nucleotide sequence operably encoding a ccdB polypeptide, wherein the first nucleotide sequence and the second nucleotide sequence are flanked within the same attR3 and attR4 sites, and wherein the polynucleotide comprises a tet Multisite having SEQ ID NO: 7.

6. The polynucleotide of claim 5 wherein the polynucleotide is selected from the group consisting of pDEST-CM1 (SEQ ID NO:5), pDEST-CM2 (SEQ ID NO:6), pDEST-CM3 (SEQ ID NO:8), pDEST-CM4 (SEQ ID NO:9), pDEST-CMZ1 (SEQ ID NO:22), and pDEST-CMZc1 (SEQ ID NO:23).

7. An expression vector comprising the polynucleotide of claim 5 .

8. A host cell comprising the expression vector of claim 7 .

9. An RNA interference (RNAi) vector comprising the polynucleotide of claim 1 .

10. An RNA interference (RNAi) vector comprising the polynucleotide of claim 5 .

11. An RNAi vector selected from the group consisting of pRIPPER-1 (SEQ ID NO:11), pRIPPER-2 (SEQ ID NO:12), pRIPPER-3 (SEQ ID NO:10), pRIPPER-4 (SEQ ID NO:13), pRIPPER-II (SEQ ID NO:14), pRIPPER-III (SEQ ID NO:15), and pRIPPER-IV (SEQ ID NO:16).

12. A method of producing one or more interfering RNA products, the method comprising expressing at least one interfering RNA product by an RNAi vector of claim 11 .

13. A host cell comprising the RNAi vector of claim 11 .

14. The RNA interference (RNAi) vector of claim 9 , wherein the RNAi vector is selected from the group consisting of pRIPPER-1 (SEQ ID NO:11), pRIPPER-2 (SEQ ID NO:12), pRIPPER-3 (SEQ ID NO:10), and pRIPPER-4 (SEQ ID NO:13).

15. The RNA interference (RNAi) vector of claim 10 , wherein the RNAi vector is selected from the group consisting of pRIPPER-II (SEQ ID NO:14), pRIPPER-III (SEQ ID NO: 15), and pRIPPER-IV (SEQ ID NO:16).

16. An RNA interference (RNAi) vector comprising a polynucleotide cassette, the polynucleotide cassette comprising a nucleotide sequence operably encoding zeomycin resistance and a nucleotide sequence operably encoding a ccdB polypeptide, wherein the polynucleotide cassette is flanked by attR1 and attR2 sites, and wherein the polynucleotide cassette comprises a G144704 cassette having SEQ ID NO: 4.

17. An RNA interference (RNAi) vector of claim 16 , wherein the RNAi vector is selected from the group consisting of pRIPPER-1 (SEQ ID NO:11), pRIPPER-2 (SEQ ID NO:12), pRIPPER-3 (SEQ ID NO:10), and pRIPPER-4 (SEQ ID NO:13).

18. The RNA interference (RNAi) vector of claim 16 , wherein the G144704 cassette having SEQ ID NO: 4 is flanked on either end by a T7 promoter and a Lac Operator sequence.

19. An RNA interference (RNAi) vector comprising a polynucleotide cassette, the polynucleotide cassette comprising a nucleotide sequence operably encoding tetracycline resistance and a nucleotide sequence operably encoding a ccdB polypeptide, wherein the polynucleotide cassette is flanked by attR3 and attR4 sites, and wherein the polynucleotide cassette flanked by attR3 and attR4 sites comprises a tet Multisite having SEQ ID NO: 7.

20. An RNA interference (RNAi) vector of claim 19 , wherein the RNAi vector is selected from the group consisting of pRIPPER-II (SEQ ID NO:14), pRIPPER-III (SEQ ID NO: 15), and pRIPPER-IV (SEQ ID NO:16).

21. The RNA interference (RNAi) vector of claim 19 , wherein the tet Multisite having SEQ ID NO: 7 is flanked on either end by a T7 promoter and a Lac Operator (LacO) sequence.