Fine Mapping Of The Main Effect QTL-AC6 And Gene Prediction For Anthocyanins Content In Kernel Of Maize.

With the improvement of living standard and the adjustment of the diet,the nutritional quality of maize is increasingly high required.Composition and content of the pigment are closely related to the nutritional quality of maize kernel.The pigment composition changes with the different colors of maize kernel.As one of the important nutritional quality,anthocyanin plays an important role in improving human health and preventing disease.Maize is not only an important raw material for the development of natural health food,but also is widely used in cosmetics,medicine and industries.Black maize is rich in anthocyanins,which possesses a variety of physiological and biochemical functions to the human body.Hence,black maize is a good material for anthocyanins study.The understanding of the genetic rule of anthocyanins provide a new reference for the application of the functional quality in natural health food,cosmetics,medicine and other fields.Meanwhile,it gives some reference for maize anthocyanins relative basic research and molecular marker assisted breeding.Black maize SDM(super dark maize)is a specific maize inbred line created by maize research institute of southwest university.In this study,SDM was used as common male parent to create two populations MoS from the cross Mo17X SDM and MuS from Mu6× SDM for studying the genetic rule of anthocyanin.QTL-AC6,the main effect QTL controlling of anthocyanin content,located in the common interval of S8-umc1014 on chromosome 6.To continue the previous research,BF4C3 was used for further positioning QTL-AC6.The progeny from one plant of the last generation of near isogenic line was used to fine mapping AC6.Then,the genome sequences of the nearby positioning area were analysised and genes within this region were annotated.The main results were as follows:1.The construction of fine genetic map of BC4F3 populationBy selecting the existing SSR polymorphism markers and the new developed polymorphism markers nearby the target QTL regions,seven pairs and eight pairs of SSR polymorphism markers were used to constract fine linkage map for MuS-BC4F3 population and MoS-BC4F3 population,respectively.The average genetic distance between each molecular marker was 0.46 cM in the fine genetic map of MuS-BC4F3 population and 1.41 cM.in MoS-BC4F3 population.2.Fine mapping of the main effect QTL-AC6 of Anthocyanins contentTwo near isogenic line segregation populations of MuS-BC4F3 and MoS-BC4F3 were used to restrict the main effect QTL-AC6 in the interval of S8-umc1014 and umc1014-mmc0523 respectively,which were close to each other,sharing the common umc1014 molecular marker.Selfing generation BC4F3 of single plant of last generation was used to fine mapping AC6.In MuS population and MoS population,10 single plants were screened respectively.According to the marker genotype and phenotype of the single plant,AC6 was defined in molecular marker ssr9-umc1014 and S73-umc1014,and the two regions overlapped.The physical distance between ssr9 and umc1014 was 255 KB,while the distance between S73 and umc1014 was 114 KB.3.Gene annotation and prediction of the positioning regionwww.ncbi.nlm.nih.gov,http://linux 1.softberry.com/berry.phtml,http://www.blast2go.de/b2ghome were used for analysis of locking region and the annotation of genes.A total of 49 transposons,29 retrotransposons,8 assumptions protein,31 gene were found.Further information comparative analysis by using NCBI(http:/blast.ncbi.nlm.nih.gov/Blast.cgi)reveals that within the positioning area,pl transcription factor was a heme oxygenase gene,participating in maize pigment regulation.From the results of other researchers who had separated related gene of anthocyanins content,pl was also located on chromosome 6.Therefore,pl transcription factor was most likely the candidate gene.The gene was not cloned with a variaty of amplification conditions in the process of gene cloning.The main reason maybe that an unkown length DNA sequence was inserted into pl transcription factor in mutant materials SDM,resulting that the gene was not cloned in SDM.