Genetic Analysis Of Genes Controlling Purple Leaf Of Lettuce And Primary Steps For The Construction Of Magic Population For Lettuce

Anthocyanin, as one natural plant pigment, has health-promoting effects. Its biosynthesis pathway and regulation have been well studied in maize, Petunia and Arabidopsis. However, little is known on anthocyanin biogenesis in lettuce. To map the gene(s) controlling lettuce purple leaf, a F2population was derived from a cross between a loose leaf lettuce with purple leaf and a stem lettuce with green leaf. A total of50F2individuals with purple leaf and50individuals with green leaf were chosen from218F2plants, and their leaves were combined to generate two pools. RNAs were extracted from the two pools and sequenced using Illumina Hi-seq. Approximate4.0Gb were obtained for each pool. Allele frequency was compared between the two pools and chromosome regions controlling lettuce leaf color were identified. The identified region was then fine mapped. The results and conclusions are as following:1. A total of166genes showed differential expression between the two pools, and6of them are from the biosynthesis pathway of anthocyanin. CAPS markers were developed for genes CHS and ANS. Linkage analysis showed that these two genes are not resposible for the the color variation between the two parents. Unfortunately, the other four genes (DFR, CHI, F3H and AT) showed no polymorphisms between the two parents. The differentially expressed genes might be genes controlling lettuce leaf color or linked with genes controlling leaf color. Alternatively, they may be differentially expressed due to the presence of anthocyanin (i.e. their expression variation is the consequence but not the cause of presence of anthocyanin).2. Analysis of RNA-seq data from the two pools showed that genes from three regions had significant difference of allele frequencies between the two pools, suggesting that the leaf color variation between the two parents is controlled by at least three genes, one in each of the three regions. They were nominated as Purple Lettuce Leaf1(PLL1), Purple Lettuce Leaf2(PLL2) and Purple Lettuce Leaf3(PLL3), respectively. PLL1and PLL2are located at approximately400Mb and100Mb from the end of short arm of chromosome05, respectively; PLL3is resided approxiately60Mb from the end of short arm of chromosome09. F2individuals, which are heterozygous at the PLL2locus but homozygous at the other two loci, were chosen to fine map the gene. Approximately6,000F2:3plants were planted and1,370green plants were chosen for further analysis. The PLL2was mapped between markers M1and M4, which are separated by-6Mb with2.85cM.3. Forty one lettuce genotypes had been selected to construct MAGIC population and24Fl hybrids were obtained. Crosses between different F1were made. These materials will be further crossed to construct MAGIC populations for GWAS analysis of lettuce.