Table I.

Synthetic oligonucleotides used

Primer NamePrimer SequenceAnnealing Temperaturea
Genotyping of hcs1 mutant
    Hcs1.for5′-CTGCCTCAGGTTCGAAACTATTAGGAC-3′
    Hcs1.rev5′-TAACACACAAAGGTAATCAACCAACC-3′
    LB35′-TAGCATCTGAATTTCATAACCAATCTCG-3′
Isolation and genotyping of hcs2 mutant
    Hcs2.Met5′-CGTTATTGAACAGTTTCGTGGTCATGC-3′
    Hcs2.Stop5′-CATCTGCGTTCACCAAAAGATGCTTC-3′
    Tag35′-CTGATACCAGACGTTGCCCGCATAA-3′
    Tag55′-CTACAAATTGCCTTTTCTTATCGACCATG-3′
    Exon75′-GGAGCTTTACTATAGGACATGGC-3′
Real-time PCR
    Hcs2.Q55′-CCAGTTGGTTCAGTTTGTGTCTCTGATATACAAC-3′60°C
    Hcs2.Q35′-GGCACTACTCGACCATCTTCCATTTCTAAT-3′60°C
    Hcs1.Q55′-CCAGTTGGTTCAGTTTGTGTCACTGATATCCAGT-3′64°C
    Hcs1.Q35′-GGCACGACTCGACCATCTTCCATTTCTAGA-3′64°C
    Left minus5′-GAACGAACTGCTTCCATTAAACTGTTGT-3′60°C
    Left plus5′-GAACGAACTGCTTCCATTAAACTATAA-3′60°C
    Right5′-AGCTCTCCTCTGCATCTGAGCATTGC-3′60°C
    Actin.55′-GTTTTGCTGGGGATGATGC-3′55°C
    Actin.35′-GGATTGAGCTTCATCGCC-3′55°C
5′-RACE experiment
    Hcs1.E45′-CCATACATTCTTTGTTCTGC-3′
BCCP2, ACC138, cloning and expression
    5′-bccp25′-AAATCTGAACATATGGCTAAAGTCTCTGG-3′
    3′-bccp25′-GCAGCTAAAGAGCTCCTTCTT-3′
    5′-acc5′-AGTAAATATCATATGGATGTAGTCC-3′
    3′-acc5′-TTATTTTGAGCTCAATCAAGATCAAGATTGGC-3′
Mutagenesis
    ATG0.for5′-CGAATAGCAGCAAGATCTTTGATTTGGCTAC-3′
    ATG0.rev5′-GTAGCCAAATCAAAGATCTTGCTGCTATTCG-3′
Cloning of HCS1 cDNA variants
    RACE.KPN5′-TTAAATTAAAGGTACCGCTCTCCTCTGCATC-3′
    STOP1.SAC5′-AAAATCTTGAGCTCATATTTTTCTTCGTCGAAC-3′
HCS1-GFP fusion cloning
    ATG1.XBA5′-GATCTAGATGGAAGCAGTTCGTTCAAC-3′
    Hcs1s.XBA5′-ACAGCTCTAGACTGCATCTGAGCATTG C-3′
    ATG2.XBA5′-CAACCTTATCTAGATTTCATCTACTG-3′
    Hcs1un.XBA5′-GTCTTCTAGATTAAAAATTGCAACTTTAAC-3′
    Hcs1.BAM5′-CCCTTAAACTGGATCCCAGTGACAC-3′
  • a The annealing temperature used for the real-time PCR is indicated.