Comparison of promoters and selectable marker genes for use in Indica rice transformation

Abstract

Key words: bar, embryogenic callus, hph, Indica riceAbstractThe effectiveness of different promoters for use in Indica rice transformation was compared. Plasmids encodingthe Escherichia coli uidA(gus) gene under the control of CaMV 35S, Emu,Act1 or Ubi1 promoters were deliveredinto cell suspension cultures by particle bombardment. Transient gene expression, 48 h after delivery, was greatestfrom plasmids utilising the constitutive promoters, Act1 and Ubi1. Gene expression in stably transformed tissuewas examined by bombarding embryogenic Indica rice calli with a pUbi1-gusplasmid and a plasmid containingeitherthe selectable markergene,hph, whichconfershygromycinresistance, orbar, whichconfersresistance to theherbicide phosphinothricin (BASTA) each under the control of the CaMV 35S, Emu, Act1 or the Ubi1 promoters.The bombardedcalli were placed on the appropriateselection media and stained for GUS activity at 1 day, 3 weeksand 5 weeks after shooting. Callus bombarded with the pUbi1-hphor the pEmu-hph constructs gave a dramaticincrease in the size of the GUS staining areas with time. No such increase in the size of GUS staining areas wasobserved in calli co-bombarded with pUbi1-gusand any of the bar containing constructs.Co-bombardment of calli with either the pEmu-hph or pUbi1-hphconstruct and a virus minor coat protein (cp)gene construct resulted in many fertile transgenic Indica rice plants, containing one to eight copies of both the hphand cp genes. These genes were stably inherited by the T