TABLE 2.
Primers used in this studya
| Name | Sequence (5′–3′) | Source of reference sequence | Position of primer in the sequence (bp) |
|---|---|---|---|
| tia_pGEM_FW | GGAGTAGTTTCGTCAGAAGT | GenBank accession no. JQ994271.1 | 4714 |
| tia_pGEM_RV | TCAATGCCTGTGTCAGTACT | GenBank accession no. JQ994271.1 | 5686 |
| FW_Kmb | GCGGACACCTGCCCCAAAACTCCATGCAAAATTGTTATCATACATGGCGATAGCTAGACT | Thermo Fisher Scientificc | 1135 |
| GenBank accession no. JQ994271.1 | 4885 | ||
| RV_Kmb | ATGAAAAAGATTATTGCGGTTTCAGCGCTTGCTGTGGTGTTCAATTCAGAAGAACTCGTC | Thermo Fisher Scientificc | 2119 |
| GenBank accession no. JQ994271.1 | 5472 | ||
| Km_check_RV | TAAAGCACGAGGAAGCGGTC | Thermo Fisher Scientificc | 2049 |
| pGEM_check_FW | CGAAAGGGGGATGTGCTGC | Promegad | 2910 |
| pGEM_check_RV | AGCGGATAACAATTTCACACAGGA | Promegad | 211 |
| tia_RT_F | TGTTCTGTGTGTCTTCCCGA | GenBank accession no. JQ994271.1 | 5109 |
| tia_RT_R | GGCGGCGATAAGAACGATAC | GenBank accession no. JQ994271.1 | 5295 |
| tia_RT_probe | GTGCGCACAGGAACATTGTA | GenBank accession no. JQ994271.1 | 5210 |
The primers designed for this study are listed. Primers used in previous studies are indicated with the corresponding reference in the text.
The nucleotides underlined in the sequence of primers FW_Km and RV_Km anneal on the plasmid pCR2.1 TOPO to amplify the Km resistance cassette, while the rest of the sequence (not underlined) anneals to the gene tia in order to amplify two regions of the gene and allow recombination (GenBank accession no. JQ994271.1).
The sequence of the plasmid pCR2.1 TOPO is available on the Thermo Fisher Scientific website, at the following address: http://tools.thermofisher.com/content/sfs/vectors/pcr2_1topo_seq.txt.
The sequence of the plasmid pGEM-T Easy is available on the Promega website, at the following address: www.promega.com/vectors/.