Research On Expression Analysis Of A Chloroplast Developing Gene GLK In Leaf Color Mutant Of Anthurium Andreaeanum L

Anthurium andreaeanum Lind., one of the most popular tropical and cut flowers,which is a perennial evergreen and herbaceous plant, belonging to Araceae. At present, target properties of Anthurium breeding are concentrated on the characteristics of spathe colors, spathe size, plant shapes (used as pot or cut flowers), leaf shapes (such as size of leaf) and so on. However, the research of leaf color mutant of Anthurium has not been reported. In2004year, one variegated plantlet, whose leaves showed half yellow and half green, resulting from chlorophyll loss of partly parenchymal tissue was obtained from the adventitious shoots regenerated from callus of Anthurium andreaeanum ’sonate’. Through the process of propagation by indirect shoot regeneration via callus or axillary bud emergence, we get a variety of mutation strains of leaf color. Through continuous screening, steady etiolated type, rubescent type and albino type were got.and got lots of propagation for experiment research. In order to select splendid plantlets from these colonies, formative mechanism and maintenance characteristics of mutant were studied. The results were as follows:1. Wild type, etiolated type, rubescent type and albino type which has a good and neat growing were used as experimental materials. Through optimized CTAB method, RNAs with high yields and high quality from Anthurium tissues. We got GLK and GAPDH cDNA fragment by reverse transcription PCR (RT-PCR) and rapidly amplication cDNA end (RACE), and analyzed the GLK gene expresstion of Anthurium wild type and various mutant.The results of real-time quantitative PCR in different phenotypes show that:GLK gene had the highest expresstion level in wild-type, Obvious higher than its expression levels in rubescent type and albinotype, its had the lowest expresstion level in rubescent type, and the expresstion in wild type was about10times of rubescent type.2. Wild type and etiolated type Anthurium with good and neat growth were used as experimental materials, and cultured for15days under the light conditions of low light10001xs, normal light50001xs and high light200001xs. Through real-time quantitative PCR of GLK gene, transmission electron microscopy, Chlorophyll and anthocyanin measure and protoplast isolation, the results show that:(1)The basic trends of GLK gene expression in Wild type and etiolated type Anthurium after the light treatment was:Normal light> low light> high light. The expression level in the normal light conditions was about2times than the high light conditions,3times than the low light conditions, and in different light conditions, the GLK gene expression in Wild type were all higher than in etiolated type.(2) There’s a large number of osmiophile globule in etiolated type Anthurium chloroplast with unclear membrane structure under low light.while wild type had fewer mitochondria and osmiophile globule, and no granulose was find in neither types. Under normal light, there’s large vacuoles in etiolated type Anthurium chloroplast with no obvious thylakoid structure, and a large number dark osmiophile globule were together observed. With clear chloroplast membrane and grana lamellas, the wild type Anthurium chloroplast had bright white oval-shaped denotes granulose and fewer osmiophile globule, while some of them expandable for contained large denotes granulose. With no normal structure, the etiolated type Anthurium chloroplast showing a cavity structure. While wild type with small denotes gaps between grana lamellas and dark grana lamellas, had lots of large osmiophile globule together.(2) The chlorophyll content under normal light condition was both higher than under low and high light conditions in wild andetiolated type Anthurium. In the same light condition, the chlorophyll content in wild type Anthurium were all higher than in the etiolated type one. The wild type had the highest chlorophyll content which was about8times of etiolated type in normal light condition.(3) Barely chloroplast could be find in protoplast of etiolated Anthurium, the protoplast of wild type had the largest number of chloroplast under nomal light condition, and least in low light condition.