Molecular cloning and sequencing of a cDNA for an auxin-repressed mRNA: correlation between fruit growth and repression of the auxin-regulated gene

Abstract

A complementary DNA (cDNA) library has been constructed in lambda gt 10 from poly(A)+ mRNA isolated from auxin-deprived strawberry receptacles. By differential plaque filter hybridization, a cDNA (lambda SAR5) to an auxin-repressed mRNA has been isolated. The expression of the auxin-repressed gene is studied at various stages of normal fruit development and in fruits of variant strawberry genotype using lambda SAR5 as a probe. Northern analyses of RNA isolated from pollinated and unpollinated fruits of various developmental stages revealed that mRNA corresponding to the lambda SAR5 clone is repressed during normal fruit development, and the level of lambda SAR5 mRNA is regulated by endogenous auxin. Furthermore, results with both normal and variant genotype strawberry fruit indicate that there is a positive correlation between growth of strawberry fruit and repression of mRNA corresponding to the lambda SAR5 clone. The lambda SAR5 cDNA has been sequenced and is 723 nucleotides in length. The deduced protein has 111 amino acid residues with a molecular mass of 12.5 kDa. The putative polypeptide starts at nucleotide position 20 and ends at 352. The molecular weight of the predicted polypeptide is in agreement with the molecular weight of the in vitro translated polypeptide of hybrid selected mRNA. A comparison of the nucleotide and deduced amino acid sequence of lambda SAR5 with nucleotide and protein sequences in data banks has not revealed any homology to known proteins.