Differential expression of genes in mandibular gland of Apis cerana nurse and forager honeybees

Abstract

Differential display PCR (DD-PCR) technique was used for detecting the differences in gene expression in mandibular glands of nurse (5-15 days old) and forager (more than 21 days old). The DD-PCR reaction was performed with 36 combinations of oligo-dT and arbitrary primers. Seventy differentially expressed bandes of nurse from forager using 19 combinations of primer were selected. Fifty (72%) out of seventy bands had higher intensity than those of forager and twenty (28%) were nurse specific genes. Moreover, in forager stages, eleven differentially expressed bands from 7 primer combinations were selected. Only one band (9%) showed higher intensity than those in nurse and ten bands (91%) were forager specific genes. Each band was reamplified and cloned into pGEM-Teasy vector. Fifty two of nurse and eleven of forager DD-PCR fragments could be successfully cloned, respectively. These cloned were selected and sequenced. The sequence analysis showed that 27 bands out of 63 contained unique sequence. Eleven genes were similar to the GenBank analysis. Two genes were found to be nurse specific genes. ATP synthase and Thioesterase were employed in order to confirm the different expression of nurse and forager using RT-PCR. The expression level of ATP synthase and Thioesterase in nurse was approximately 1.78 and 1.40 times higher than those of forager, respectively. After the nucleotide sequences were analyzed, it was found that from each 36 DD-PCR bands contained more than one cDNA type. When compared the nucleotide sequences with GenBank database, 21 genes were found similar to Apis spp. Eight of them were nurse specific genes.