Table 3.
PCR primers used to generate the cassettes for deleting STL1 , GUT1, and GUT2 in CBS 6412-13A and for the PCR-based verification of the respective deletions
| Gene | Primer | Sequence (5′-3′) |
|---|---|---|
|
STL1 |
astl1-loxP-fw |
CACTCATAGTATATAAACAAGCCCTTTATTGATTTTGAATAATTAcagctgaagcttcgtacgc |
| |
astl1-loxP-rv |
TCAAAGCCCTCTGAAGATTTTGGGACCTGCCTCTGGAGAACAAACgcataggccactagtggatctg |
| |
bverif-stl1∆-fw |
TGGTTCACCTTTGATAGGGC |
| |
bverif-stl1∆-rv |
TGAAACTGCTTGACCTGTGG |
|
GUT1 |
agut1-loxP-fw |
TGTGGGGGGATGCCTGTTCTCGAACCATATAAAATATACCATGTGcagctgaagcttcgtacgc |
| |
agut1-loxP-rv |
CTAGATCTCGCAGTACTGTTTTTGGCGAATCGTGTAGCTTTTCCAgcataggccactagtggatctg |
| |
bverif-gut1∆-fw |
GTGTGGAGTAGCATAGTGAGG |
| |
bverif-gut1∆-rv |
AATGCTAGAGTCGTCAGTGCG |
|
GUT2 |
agut2-loxP-fw |
GTCTAAAGCAAGGACTCTCCCTCCCTTATCTTGACCGTGCTATTGcagctgaagcttcgtacgc |
| |
agut2-loxP-rv |
TTGCAAAATGGCGTCACTGTATGGGCCCGTGGCATTGACCACACAgcataggccactagtggatctg |
| |
bverif-gut2∆-fw |
TCCGATACGTTATCCACCCAA |
| bverif-gut2∆-rv | TTCCTCAGCCATTTGTCTGT |
aGene deletion cassettes; sequences complementary to the loxP regions within the disruption cassettes are shown in lower case.
bVerification of gene deletions.