TABLE 1.
PCR primers used in this study and their products
| Primer combination | Primer sequences (5′→3′) | Product | Amplicon length(s) (bp) |
|---|---|---|---|
| 1 | TACTTGGTGGATTAGAAGCAATGTCAACTTCAAATCTTTTCAC + GCCATCATGACACCATTTGA | easA with promoter extensiona | 1,841 |
| 2 | GTGAAAAGATTTGAAGTTGACATTGCTTCTAATCCACCAAGTA + GGCTTGGATTGAACGGTCATGGTGCGGAGTGCCTACTCTA | Promoter with easA and easH extensions | 830 |
| 3 | TAGAGTAGGCACTCCGCACCATGACCGTTCAATCCAAGCC + CCTAGCTATCCATGCTCAAGC | easH with promoter extension | 1,135 |
| 4 | CGTATCACCGAGACAAAGAGG + TTGGCCATCACCTAACTATCTTG | easA-promoter-easH fusion construct | 3,250 |
| 5 | GTGAAAAGATTTGAAGTTGACATTGCTTCTAATCCACCAAGTA + GGATAACCATGGTAATATCATGGTGCGGAGTGCCTACTCTATAG | Promoter with easA and cloA extensions | 831 |
| 6 | CTATAGAGTAGGCACTCCGCACCATGATATTACCATGGTTATCC + AACACGCTAAGGGCAACAAG | cloA with promoter extension | 2,746 |
| 7 | CGTATCACCGAGACAAAGAGG + GCAACAAGCGATAAGCGTTAG | easA-promoter-cloA fusion construct | 4,905 |
| 8 | GCGAATGGATTTGATCTCGT + CCAGCGAGAGTTAGCAAGGT | easA internal sequences | 447 |
| 9 | CCAACGGTTCTCCCTTACTTC + GCACTATCTTGCCGCTCAGT | easH internal sequences | 611 |
| 10 | TTCCCGGCACGAGCTTTGCG + CTTAGAGTGCACCTCAGACGAC | cloA internal sequences | 296 (cDNA), 362 (gDNAb) |
a
Extension refers to incorporation of an additional 20 to 23 nucleotides at the 5′ end of a primer to add sequences that will facilitate a later fusion PCR.
b
gDNA, genomic DNA.