Abstrakt

The Selection of Reference Genes for a Comparison of the Biosynthesis of Flavoids from L. confusa and L. japonica

Qiao-cheng MO, Mu-xiu TAN, Dan-na HUANG, Feng-hua SHI*

stability of 8 candidate reference genes in 4 tissues of L. confusa and L. japonica were evaluated by using four algorithms to assess the validity of reference genes for their use in accurate normalization of qRTPCR data. The 8 candidate reference genes are β-TUB, eIF-4a, ACT11, eEF-1α, GAPDH, UBQ10, 18S and 25S, and the 4 tissues were leaves, green buds, white buds and white flowers. Results showed that the most stable genes selected by the four algorithms of these 8 genes are slightly different for these tissues, but the least stable genes were the same in each case. ACT11, eIF-4a and β-TUB are the three most stable genes as selected by geNorm, β-TUB is the most stable as judged by Delta CT and Normfinder, and GAPDH by Bestkeeper. 18S and 25S were the two least stable genes as judged by all the four algorithms. To verify the validity of these selected genes by the four methods, the comparative expression level of FSâ�?¡ gene in these 8 tissues were normalized with them. Similar expression profiles of FSâ�?¡ were seen with the most stable genes selected by the four methods and the highest was found in the leaves and the lowest in the flowers of L. confusa. But they are apparently different from the profiles when subjected to normalization by the least two stable genes: 18S and 25S. This demonstrated that apporatative reference genes are indispensable for gene profile assays, and these recommended genes by the four algorithms are all reliable. This is the first report on the selection of candidate reference genes in Lonicera species and this will provide a more precise comparsion of gene expression in the biosynthesis pathway of flavoids between L. confuse and L. japonica.