Polymerase chain reaction species diagnostic assay for Anopheles quadrimaculatus cryptic species (Diptera: Culicidae) based on ribosomal DNA ITS2 sequences.
Species-specific differences in the nucleotide sequences of the 2nd internal transcribed spacer (ITS2) of nuclear ribosomal DNA (rDNA) were used to develop a diagnostic assay based on the polymerase chain reaction (PCR) that can distinguish 4 of the 5 cryptic sibling species in the common malaria mosquito, Anopheles quadrimaculatus Say, complex. The assay requires only a small amount of tissue from an individual mosquito and a mixture of 5 PCR primers. The plus strand universal primer is derived from a sequence in the 5.8S coding region that is identical in all members of the complex. The 4 minus strand primers were selected from species-unique sequences within the ITS2 region. PCR amplification produces a different sized fragment for each of the 4 species which can be visualized readily under ultraviolet light after electrophoresis through an ethidium bromide-containing agarose gel. The assay has been developed and tested only with An. quadrimaculatus complex specimens from Florida populations.
Cornel
A J
AJ
Entomology Branch, Division of Parasitic Diseases, Centers for Disease Control and Prevention, Chamblee, GA 30341, USA.
Porter
C H
CH
Collins
F H
FH
D000818:Animals; D000852:Anopheles / Q000145:classification* / Q000235:genetics*; D001483:Base Sequence; D017931:DNA Primers; D004275:DNA, Ribosomal / Q000235:genetics*; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D016415:Sequence Alignment
D017931:DNA Primers; D004275:DNA, Ribosomal