Three β-galactosidase cDNA clones related to fruit ripening in papaya (Carica papaya)

Abstract

β-Galactosidase (EC 3.2.1.23) is a hydrolase which plays an important role in cell wall modification and fruit softening during ripening. In this study, three full-length β-galactosidase cDNA clones were successfully obtained from papaya mesocarp using different approaches. pPGBII which is 2,771 bp in size, was isolated from a papaya ripe mesocarp cDNA library using a heterologous probe. The other two cDNA clones, pBG(a) and pBG(b), which are 3,168 and 2,580 bp in size, respectively, were amplified from ripe papaya fruit using the reverse transcriptase polymerase chain reaction (RT-PCR) and rapid amplification of cDNA end (RACE) approaches. The pPBGII, pBG(a) and pBG(b) cDNAs, respectively, were expected to yield a putative mature polypeptide of 79, 91 and 56 kDa with isoelectric points (pI) of 8.12, 8.75 and 8.26. The genomic DNA gel blot analysis indicated that all of the β-galactosidase genes exist as multiple copies in the papaya genome hence they belong to a multigene family. All three cDNA clones were expressed in fruits during ripening with varying patterns. In mesocarp, pPBGII mRNA was only expressed during ripening and peaked at the half-ripe stage when the fruits undergo dramatic softening. Meanwhile, pBG(a) mRNA expression increased from the immature green stage to the half-ripe stage where it reached maximum level before declining. pBG(b) mRNA level accumulated abundantly at the mature green stage and decreased thereafter. Therefore, we suggest that pPBGII and pBG(a) cDNA clones characterized in this work may be involved in fruit softening during papaya ripening while the fruit-specific pBG(b) may be related to early ripening stage.