Molecular cloning of the complete 11S seed storage protein gene of Coffea arabica and promoter analysis in transgenic tobacco plants

• Published in: Plant physiology and biochemistry Vol. 37; no. 4; pp. 273 - 282
• Main Authors: Marraccini, Pierre, Deshayes, Alain, Pétiard, Vincent, Rogers, William John
• Format: Journal Article
• Language: English
• Published: Paris Elsevier Masson SAS 01.04.1999; Elsevier
• Subjects: 11S storage protein; Agronomy. Soil science and plant productions; Biological and medical sciences; Classical and quantitative genetics. Population genetics. Molecular genetics; Coffea arabica; coffee genetic engineering; endosperm–specific promoter; Fundamental and applied biological sciences. Psychology; Generalities. Genetics. Plant material; Genetics and breeding of economic plants; Molecular genetics; CSPD, disodium 3-(4-methoxyspiro{1,2-dioxetane-3,2'-(5'-chloro)tricyclo[3.3.1.1 37]decan}-4-yl) phenyl phosphate; Coffea arabica; TMAC, tetramethyl ammonium chloride; GUS, β -glucuronidase; UTR, untranslated region; csp1 , coffee storage protein gene; coffee genetic engineering; 11S storage protein; CaMV, cauliflower mosaic virus; WAF, weeks after flowering; IPCR, inverse polymerase chain reaction; endosperm–specific promoter; Ta, annealing temperature; Storage protein; Nucleotide sequence; Transcription promoter; Tissue specificity; Stimulant plant; Gene expression; Nicotiana tabacum; Regulation(control); Gene; Dicotyledones; Angiospermae; Rubiaceae; Spermatophyta; Endosperm; Transgenic plant; Solanaceae; Molecular cloning
• ISSN: 0981-9428; 1873-2690
• DOI: 10.1016/S0981-9428(99)80025-4

In this paper, we present the complete nucleotide sequence of the csp1 gene from Coffea arabica coding for the 11S-globulin seed storage protein. To investigate the sequences responsible for the regulated expression of this seed-specific coffee storage protein gene, about 1 kb of the 5'-upstream region from the csp1 gene was isolated using inverse polymerase chain reaction (IPCR) and then sequenced. Several DNA boxes were found in this coffee sequence that had similarity to those previously identified as being essential for grain (endosperm) specific expression in other plants. To study the ability of this sequence to direct grain-specific expression, the whole fragment, as well as a series of 5' deletions, was fused to the reporter gene β-glucuronidase ( uidA) and analysed in transgenic Nicotiana tabacum plants. GUS measurements showed that all the deletions of the csp1 promoter directed the expression of the reporter gene in tobacco grain but not in the other tissues examined. GUS activities also revealed that the csp1 promoter constructs function as very strong promoters by comparison to the strength of the cauliflower mosaic virus (CaMV) 35S promoter. Therefore, this 11S promoter could represent a useful tool to change the expression of targeted genes in the grain of transgenic coffee plants.