Map-based Cloning And Expression Analysis Of C (Chromogen ForAnthocyanin) Gene In Rice (Oryza Sativa L.)

Anthocyanin is widely present in different plant tissues, such as apiculus, stigma, sterile lemma,leaf sheath, leaf blade, pericarp and shell, and it is the basis of the production of different kinds of color.The tissue-specific pigmentation has been used as the important agronomic traits for thevariety identification and linkage analysis in rice. Especially, the apiculus and stigma colorations are notonly the important morphological markers for variety identification and purification, but alsoconducive to studying phenotypic and genetic diversity during the domestication process.Yunnan japonica landrace LTH exhibites red apiculus and purple stigma, while indica cultivars93-11and Longhong exhibites achromatic apiculus and stigma. F1hybrids derived from the intercrossesof LTH with93-11and longhong displayed red apiculus and purple sitgma, and both of F2populationssegregated into either red or achromatic apiculus and either purple or achromatic stigma, and thephenotypic segregation ftted3:1ratio in each F2populations. The apiculus coloration matched tostigma coloration in each F2plant, suggesting that the red apiculus and purple stigma trait of LTH wascontrolled by a pair of dominant genes.Using1851F2recessive individuals derived from the crosses of LTH/93-11and LTH/Longhong,the gene governing red apiculus and purple stigma was mapped within a70.8kb interval between twomolecular markers dCAPS13and RM19561, co-segregating with the marker C3C8. There werefourteen recognizable open reading frames (ORFs) within this targeting region, of which a homolog ofanthocyanin regulatory C1gene LOC_Os06g10350, was likely the candidate gene for the targeting gene.The sequences of LOC_Os06g10350were sequenced and compared among LTH,93-11and Longhong,showing that LOC_Os06g10350in93-11and longhong was different from that in LTH by a10bpdeletion, which was located in the third exon795position and may cause a frameshift mutation andpremature translation.We construced complementation and overexpression vectors carrying LOC_Os06g10350andintroduced them into the corresponding receptors. The result indicated that the Kitaake transgenic T0and T1plants exhibited red apiculus as LTH, and with the maturity of seeds, the apiculus colorationgradually deepened, but the stigma coloration was achromatic. Therefore, LOC_Os06g10350wasverified to be the OsC gene.OsC was expressed in various organs, including young leaf, stem, leaf blade, leaf sheath and youngpanicle, with a signifcantly high level of expression detected in leaf blade and leaf sheath. OsC encodeda R2R3-MYB protein that activated the transcription of the anthocyanin structural genes. The OsC geneexhibited high homology to C1and pl from Zea mays and involved in regulation of anthocyanin in ricetissues. These results will be helpful for dissecting the function of OsC and the regulatory mechanism ofOsC in biosynthetic pathway of anthocyanins.