Table 1.
Comparison of the Gateway Cloning System for the Current Human ORFeome Project to the Previous C. elegans ORFeome Project (Reboul et al. 2003)
|
|
C. elegans ORFeome |
Human ORFeome |
|---|---|---|
| PCR DNA Template | cDNA library | isloated cDNA clone |
| Number of PCR Cycles | 35 | 20 |
| Taq Polymerase | Platinum Taq DNA Polymerase High Fidelity | KOD Hot Start DNA polymerase |
| Mutation Rate | 1/1500 | 1/35,000 |
| AttB1 Site Sequence | B1:acaaGtttgtacaaaaaagCAggct | B1.1: acaaCtttgtacaaaaaagTTG |
| AttB2 Site Sequence | B2: acCactttgtacaagaaagCtgggt | B2.1: acAactttgtacaagaaagTtg |
| pDONR Vector | pDONR201 | pDONR223 |
| 5′ ORF Specific Primer | 5′-ggggacaagtttgtacaaaaaagcaggcttg(nnn)x | 5′-ggggacaactttgtacaaaaaagttggcatg(nnn)x |
| 3′ ORF Specific Primer
|
5′-ggggaccactttgtacaagaaagctgggta(nnn)x
|
5′-ggggacaactttgtacaagaaagttggg(t or c)a(nnn)x
|
