Screening molluscan cDNA expression libraries with anti-shell matrix antibodies.
In a previous paper [Marin et al., Protein Expr. Purif. 23 (2001) 175], we showed that polyclonal antibodies raised against molluscan shell matrices could be useful tools for visualizing shell proteins after a preparative fractionation of the shell matrix. In this paper, we have used the same antibodies for screening a cDNA library constructed from mantle tissues of the nacro-prismatic bivalve Pinna nobilis. The immunoscreening led to the identification of a new protein, mucoperlin [Marin et al., J. Biol. Chem. 275 (2000) 20667], which was subsequently overexpressed. A polyclonal antibody was obtained from the recombinant mucoperlin. In a control assay, we unambiguously demonstrated that this antibody and one of the sera used for the initial screening hybridize with the same clones. We assess that screening cDNA libraries with antibodies elicited against unfractionated calcifying matrices is a good alternative to oligonucleotide screening techniques, particularly in the field of molluscan biomineralization where only few gene sequences are known.
Marin
Frédéric
F
IsoTis NV, Prof. Bronkhorstlaan 10, Gebouw D 3723, MB Bilthoven, The Netherlands. frederic.marin@u-bourgogne.fr
de Groot
Klaas
K
Westbroek
Peter
P
D000818:Animals; D015153:Blotting, Western; D018076:DNA, Complementary / Q000235:genetics; D015870:Gene Expression; D015723:Gene Library; D007106:Immune Sera / Q000276:immunology*; D008974:Mollusca / Q000235:genetics*; D009077:Mucins / Q000235:genetics* / Q000276:immunology* / Q000302:isolation & purification; D011994:Recombinant Proteins / Q000235:genetics / Q000276:immunology
D018076:DNA, Complementary; D007106:Immune Sera; D009077:Mucins; D011994:Recombinant Proteins; C411849:mucoperlin