1. Establishment Of An Efficient Auxin Biosensor Marker System And Its Application In Rice 2. Genetic Diversity Analysis Of Human UGT1 Gene Family

Auxin,as one dispensable plant hormone,plays diversified and important roles in the process of plant morphogenesis,such as embryogenesis,vascular development and root formation.The precise locations and patterning mechanisms of auxin have been reported in several plant species,but remain obscure in monocot model plant rice（Oryza sativa）.Furthermore,even less is known about auxin detailed functions in rice development,on account of the very limited number of auxin-related rice mutants identified till now.Considering the shortcomings of DR5-GUS report lines,which include the low spatial and temporal resolution of GUS signals,and stringent requirements for the experimental conditions,we choose to establish a stable and efficient auxin marker system by precisely revealing auxin accumulation sites at the cellular level.This system adopted DR5-VENUS in combination with DII-VENUS construct which is based on the classic SCF^TIR1-dependent auxin signaling pathway,and driven by Maize Ubiquitin-1 promoter.This research for the first time introduces a set of effective auxin tools into rice,which offers a feasible method of following auxin activities at the cellular level,and provides key clues for illustrating the relationship between auxin and rice flower formation.Auxin accumulation sites were observed at various rice tissues and developmental processes by live imaging.Stimulating transgenic lines with chemicals,hormones and gravity showed these auxin sensors were quite sensitive to1-Naphthaleneacetic acid（NAA）and N-1-Naphthylphthalamic acid（NPA）.Furthermore,auxin signal redistribution observed in vivo confirmed the crosstalk between auxin and cytokinin Trans-zeatin（TZ）,and also auxin rapid response to gravity,which were further confirmed by qRT-PCR results.Four pin-formed 1（PIN1）proteins were found in rice,and through protein sequence alignment and phylogenetic analysis,we found that OsPIN1a and OsPIN1b are orthologous to Arabidopsis PIN1,while OsPIN1c,OsPIN1d and AtPIN1 gene are paralogous.Topological prediction showed that rice PIN1s have similar transmembrane structures.During stages of spikelet development,immunostaining analyses using PIN1 antibody,and the confocal observation of ProOsAUX1:OsAUX1-sGFP double labelling system both revealed highly overlapping expression patterns between PIN1,auxin resistant 1（OsAUX1）and DR5-VENUS,providing the strong evidence that hormone auxin is indeed essential for rice flower formation.Taken together,we successfully developed two auxin reporter systems DR5-VENUS and DII-VENUS in rice,and validated their reliability and sensitivity.Using these two reporters,we revealed responsive sites,quantitative levels and dynamic changes of auxin during rice development,further discovering auxin potential funtional differentiation at shoot apical meristem（SAM）,lateral root emergence and inflorescence branching,by comparing auxin distribution in diocot and monocot model plants.Besides we analyzed the expression pattern of rice auxin-related OsAUX1 gene,and found that this gene is mainly expressed at the central vasculature tissue,epidermis of meristematic zone and differentiation zone in primary root.Abnormal gravitropic response,altered root angle and shortened root hair phenotypes were observed in osaux1 mutants.The application of the established auxin reporter system and GC-MS quantification results further revealed that auxin response and level were obviously elevated under low phosphorus（P）treatment,which indicate that auxin is involved in regulating root hair elongation by OsAUX1 protein transport for plant adaptive growth in the low P environment.This work provides new molecular tools for the rice auxin research,and uncovers the strong link between auxin and agronomical traits such as yield and root architecture;moreover firstly validate the novel mechanism that OsAUX1 can manipulate root system for controlling response to outside P stimuli.Human adaptive immune and defense system requires massive amount of biological molecules inside to defend against the numerous bacteria,virus and chemicals in the huge living environment.The limited size of human genome evolves out kinds of regulating mechanisms,such as gene duplication,gene transposition,gene conversion,DNA rearrangement and copy number variation,which provide the strong genetic basis for meeting survival demands of human body.Products encoded by UDP-glucuronosyltransferase family 1（UGT1）belong to Class II drug metabolism enzymes.By transferring the glucuronic acid（GA）residue of uridine diphosphate glucuronic acid（UDPGA）donor to receptors through glucuronidation,toxic substances in vivo or in vitro（environmental toxics or therapeutic drugs）are excreted out of human body after transforming them into hydrophilic molecules.Beside alternative splicing and promoter mechanisms,single nucleotide polymorphism（SNP）can also affect UGT1 enzyme activities as reported,some specific sites among which are related with certain clinical diseases,such as hyperbilirubinemia,Gilbert syndrome and side effects of anti-cancer drug irinotecan.Along with adding the diagnosis of some pathogenic SNP sites of UGT1 in clinical application of some populations in the world,it is urgent to find out the status of UGT1 SNP in Chinese group.This research firstly illustrates the UGT1 SNP sites,population frequencies,linkage levels,and also selects out biomarkers for different structure levels,which is essential for UGT1 clinical diagnosis in the near future in China.In this research,through the large-scale resequencing data of 253 humangenomic DNA samples from the peripheral blood,we systematically analyzed molecular traits of SNPs at the UGT1 gene locus,covering nine first and five constant exons,introns and promoters,about 17.7 kb in total.We found 101 SNPs in this gene cluster,among which 15 SNPs are novel,computed allele frequencies in this population,and on the basis of these we constructed Chinese ethnic-specific linkage disequilibrium（LD）map with HAPLOVIEW.Furthermore we inferred the potential existing haplotypes by using Bayesian statistical method,and further selected out tag SNPs with STAMPA program for telling different haplotypes,which lay basis for SNP genotyping in UGT1 clinical pharmacogenomics.At last,through comparison among four haplotype map（Hap Map）populations（Japanese,JPT;Caucasian,CEU;African,YRI）,we pinpointed the Chinese ethnic-specific genetic traits of molecular diversity.This research provides a solid theoretical foundation for UGT1 gene association anlysis and future pharmacogenomics,but also have profound guiding significance for clinicnal disease risk assessment and personal therapy in China.