Sequencing and characterization of a cDNA encoding a ferritin subunit of Colorado potato beetle, Leptinotarsa decemlineata.
A differentially expressed cDNA fragment (P311) from Colorado potato beetle (CPB), Leptinotarsa decemlineata (Say), was identified by restriction fragment differential display-polymerase chain reaction (RFDD-PCR) technique, and showed a strong similarity to ferritin heavy chain subunits of other organisms. Based on P311, we constructed specific primers and obtained a 840-bp cDNA fragment spanning the open reading frame of CPB ferritin subunit using the rapid amplification of cDNA ends (RACE) technique. The sequence encodes 213 amino acid residues, including a 19 amino acid signal peptide. The sequence has a conserved cysteine in the N-terminus and has the seven conserved residues that comprise the ferroxidase center, which is the feature of heavy chain ferritins of vertebrates. The CPB ferritin subunit has high amino acid sequence identity with the Apriona germari (69.3%), Galleria mellonela (54.5%), Manduca sexta (54.0%), Drosophila melanogaster (53.2%), Calpodes ethlius (51.4%), and Nilaparvata lugens (47.6%) but lower identity with the Anopheles gambiae (38.7%) and Aedes aegypti (37.8%). Using Northern blot analysis, the subunit mRNA was identified from fat body and midgut of 4th instars with much higher mRNA levels found in midgut than that in fat body (2.5-fold). Nevertheless, only the levels of mRNA in fat body was induced by dexamethasone (1.5-fold).
Qiu
Lihong
L
Department of Veterinary and Animal Sciences, University of Massachusetts, Amherst, MA 01003, USA.
Gao
Jian-Rong
JR
Clark
J Marshall
JM
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D015152:Blotting, Northern; D016000:Cluster Analysis; D001517:Coleoptera / Q000235:genetics*; D017931:DNA Primers; D018076:DNA, Complementary / Q000235:genetics; D003907:Dexamethasone; D005293:Ferritins / Q000235:genetics*; D008969:Molecular Sequence Data; D010802:Phylogeny; D016133:Polymerase Chain Reaction / Q000379:methods; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D017385:Sequence Homology
D017931:DNA Primers; D018076:DNA, Complementary; D003907:Dexamethasone; D005293:Ferritins