ABSTRACT
OBJECTIVE: Based on the system of regeneration,the genetic transformation system of Lonicera macranthoides was established. METHODS: Tissue culture method of seedlings, Agrobacterium tumefaciens mediated transformation method of explants, report gene was detected by gus staining and PCR. RESULTS: The efficient transformation time was 8 minutes of infection. The good transformation rate was gained with the kanamycin 35 mg/L and cefotaxime 600 mg/L. The concentration of kanamycin had a leading effect on bud differentiation between two antibiotics, and bud induction rate reached extremely significant difference. Results of gus staining and PCR proved that the gus gene was integrated into Lonicera macranthoides genome. CONCLUSION: The genetic transformation system of Lonicera macranthoides leaves mediated by Agrobacterium tumefaciens EHAlO5 was established for the first time.
Subject(s)
Agrobacterium tumefaciens/genetics , Lonicera/genetics , Plants, Medicinal/genetics , Transformation, Genetic , Cefotaxime/pharmacology , Genetic Vectors , Kanamycin/pharmacology , Lonicera/growth & development , Plant Leaves/genetics , Plant Leaves/growth & development , Plants, Genetically Modified/genetics , Plants, Genetically Modified/growth & development , Plants, Medicinal/growth & development , Polymerase Chain Reaction , Tissue Culture Techniques/methodsABSTRACT
BACKGROUND: A three-generation Chinese family with Hailey-Hailey disease (HHD) was identified and characterized. The proband developed HHD with severe recurrent blisters and crusted erosions involving the body folds. Skin biopsy studies showed epidermal hyperkeratosis and defects in cell-to-cell adhesion. Three other members in the family were also affected with HHD and had the same clinical manifestations. The purpose of this study was to identify the pathogenic gene or mutation in the family. METHODS: All exons and exon-intron boundaries of ATP2C1 were polymerase chain reaction (PCR) amplified and sequenced with DNA samples from the proband. Restriction fragment length polymorphism (RFLP) analysis for the intron 23-exon 24 boundary of ATP2C1 was performed in all family members and in 100 normal control subjects. RESULTS: A novel 2-bp deletion (c.2251delGT) was detected in exon 24 of the ATP2C1 gene. The mutation was present in the three other affected family members and in two asymptomatic young carriers, but not in the other normal family members or the 100 normal controls. The mutation resulted in a frameshift change and led to the formation of a premature termination codon (PTC) four amino acid residues downstream from the sixth transmembrane domain. CONCLUSIONS: Our results indicate that the novel c.2251delGT (p.V751fs) mutation in the ATP2C1 gene is responsible for HHD in this Chinese family. This study expands the spectrum of ATP2C1 mutations associated with HHD.
