Introduction 2001 2004 1995 1998 2000 1995 1996 2000 2005 1995 2006 Arabidopsis 2001 2004 1994 1997 1986 1994 2000 2001 2002 2005 1997 2007a b 1997 2001 1997 2000 Arabidopsis thaliana 2001 2000 1998 Arabidopsis 1998 2006 AtMYB60 2007a AtMYB60 DFR Arabidopsis Materials and methods Plant materials and growth condition To investigate the effects of exogenous AtMYB proteins on anthocyanin biosynthesis in lettuce leaves at the molecular level, Jinjachuckmyun plants, supplied by Syngenta (Seoul, Korea), were used. The plants were grown at the National Institute of Agricultural Biotechnology in Suwon, South Korea. Fresh lettuce leaves were plucked from the plants, frozen immediately in liquid nitrogen and stored at −80°C until required. AtMYB Arabidopsis AtMYB4 AtMYB29 AtMYB30 AtMYB34 AtMYB51 AtMYB60 Vector construction AtMYB AtMYB Mlu Xba AtMYB4 AtMYB29 AtMYB30 AtMYB34 AtMYB51 AtMYB60 Mlu Xba AtMYB 2 Agrobacterium tumefaciens AtMYB Agrobacterium −2 −1 A. tumefaciens −1 −1 −1 −1 −1 −1 −1 −1 −1 −1 −1 −1 Anthocyanin extraction and HPLC analysis Anthocyanin in the lettuce plants was quantified by HPLC at a detection wavelength of 510 nm, comparing the sample retention times and peaks with those of known standards. The relative quantification of each phenolic compound samples was expressed as the equivalent quantity of purified standards for each treatment. −1 RT-PCR analysis AtMYB4 AtMYB60 AtMYB4 AtMYB60 CHS F3H DFR UFGT CHS F3H DFR UFGT rRNA bar CHS F3H DFR UFGT rRNA bar Results Arabidopsis Arabidopsis AtMYB AtMYB 1999 Arabidopsis 1998 Arabidopsis AtMYB 1998 AtMYB4 AtMYB29 AtMYB30 AtMYB34 AtMYB51 AtMYB60 1 Fig. 1 black gray line AtMYB4 AtMYB29 AtMYB30 AtMYB34 AtMYB51 AtMYB60  AtMYB AtMYB Agrobacterium tumefaciens 2 AtMYB Fig. 2 LB RB P35S PolyA Tnos  AtMYB4 AtMYB29 AtMYB30 AtMYB34 AtMYB51 AtMYB4 AtMYB60 3 bar bar AtMYB4 AtMYB60 AtMYB4 AtMYB60 Fig. 3 AtMYB4 AtMYB60 Bar WT  AtMYB 2007a AtMYB A. tumefaciens AtMYB60 4 Fig. 4 Arabidopsis MYB AtMYB60 a WT AtMYB60 b AtMYB  AtMYB60 AtMYB 2007a AtMYB29 AtMYB30 AtMYB34 AtMYB51 AtMYB60 5 AtMYB60 Fig. 5 WT AtMYB Peak C D  RNA expression profiles of anthocyanin biosynthetic enzymes suggest their coordinated regulation in AtMYB60 transgenic lettuce plants 4 6 2007a 2007a AtMYB4 AtMYB60 Fig. 6 AtMYB4- AtMYB60 CHS F3H DFR UFGT  6 CHS F3H UFGT DFR Discussion AtMYB 1998 1998 2007 1996 CHS 1997 2007a The lettuce has merit as a model plant system to further characterize the functions of those gene products in anthocyanin metabolism, because of its leaf color is red under field conditions and green under chamber condition when exposed to UV-B irradiation. The findings described in this study further validate the use of lettuce as a model plant because it offers experimental versatility in terms of its genetic regulation of the inhibition of leaf pigmentation. The function of the MYB proteins as activators (green to red in a growth chamber) or repressors (red to green in the field) of anthocyanin biosynthesis can be characterized directly from a phenotypic analysis of the lettuce leaf color. This system has potential applications as a research tool in a number of areas of plant molecular biology. AtMYB60 4 AtMYB60 AtMYB60 AtMYB60 AtMYB60 1994 1997 2005 AtMYB60 AtMYB60 DFR AtMYB60 DFR 7 Fig. 7 AtMYB60  AtMYB60 DFR 1999 Arabidopsis 2000 2005 CHS DFR LDOX 1995 2003 2001 2003 AtMYB60 AtMYB60 2005 AtMYB60 2005 AtMYB60 DFR