{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,6]],"date-time":"2026-10-06T04:17:21Z","timestamp":1791260241516,"version":"4.1.0"},"reference-count":21,"publisher":"American Association for the Advancement of Science (AAAS)","issue":"5451","content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Science"],"published-print":{"date-parts":[[2000,1,14]]},"abstract":"<jats:p>\n                    Rice (\n                    <jats:italic>Oryza sativa<\/jats:italic>\n                    ), a major staple food, is usually milled to remove the oil-rich aleurone layer that turns rancid upon storage, especially in tropical areas. The remaining edible part of rice grains, the endosperm, lacks several essential nutrients, such as provitamin A. Thus, predominant rice consumption promotes vitamin A deficiency, a serious public health problem in at least 26 countries, including highly populated areas of Asia, Africa, and Latin America. Recombinant DNA technology was used to improve its nutritional value in this respect. A combination of transgenes enabled biosynthesis of provitamin A in the endosperm.\n                  <\/jats:p>","DOI":"10.1126\/science.287.5451.303","type":"journal-article","created":{"date-parts":[[2002,7,27]],"date-time":"2002-07-27T05:50:14Z","timestamp":1027749014000},"page":"303-305","source":"Crossref","is-referenced-by-count":1840,"title":["Engineering the Provitamin A (\u03b2-Carotene) Biosynthetic Pathway into (Carotenoid-Free) Rice Endosperm"],"prefix":"10.1126","volume":"287","author":[{"given":"Xudong","family":"Ye","sequence":"first","affiliation":[{"name":"Institute for Plant Sciences, Swiss Federal Institute of Technology, CH-8092 Zurich, Switzerland."}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Salim","family":"Al-Babili","sequence":"additional","affiliation":[{"name":"University of Freiburg, Center for Applied Biosciences, D-79104 Freiburg, Germany."}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Andreas","family":"Klo\u0308ti","sequence":"additional","affiliation":[{"name":"Institute for Plant Sciences, Swiss Federal Institute of Technology, CH-8092 Zurich, Switzerland."}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jing","family":"Zhang","sequence":"additional","affiliation":[{"name":"Institute for Plant Sciences, Swiss Federal Institute of Technology, CH-8092 Zurich, Switzerland."}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Paola","family":"Lucca","sequence":"additional","affiliation":[{"name":"Institute for Plant Sciences, Swiss Federal Institute of Technology, CH-8092 Zurich, Switzerland."}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Peter","family":"Beyer","sequence":"additional","affiliation":[{"name":"University of Freiburg, Center for Applied Biosciences, D-79104 Freiburg, Germany."}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ingo","family":"Potrykus","sequence":"additional","affiliation":[{"name":"Institute for Plant Sciences, Swiss Federal Institute of Technology, CH-8092 Zurich, Switzerland."}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"221","reference":[{"key":"e_1_3_1_2_2","doi-asserted-by":"crossref","first-page":"96","DOI":"10.1093\/jn\/119.1.96","volume":"119","author":"Sommer A.","year":"1988","unstructured":"Sommer A., J. Nutr. 119, 96 (1988).","journal-title":"J. Nutr."},{"key":"e_1_3_1_3_2","unstructured":"J. P. Grant The State of the World's Children (Oxford Univ. Press Oxford 1991)."},{"key":"e_1_3_1_4_2","doi-asserted-by":"crossref","first-page":"63","DOI":"10.1146\/annurev.nu.09.070189.000431","volume":"9","author":"West K. P.","year":"1989","unstructured":"West K. P., Howard G. R., Sommer A., Annu. Rev. Nutr. 9, 63 (1989).","journal-title":"Annu. Rev. Nutr."},{"key":"e_1_3_1_5_2","first-page":"225","volume":"70","author":"Humphrey J. H.","year":"1992","unstructured":"Humphrey J. H., West K. P., Sommer A., WHO Bull. 70, 225 (1992).","journal-title":"WHO Bull."},{"key":"e_1_3_1_6_2","doi-asserted-by":"crossref","first-page":"53","DOI":"10.1079\/PNS19830007","volume":"42","author":"Pirie A.","year":"1983","unstructured":"Pirie A., Proc. Nutr. Soc. 42, 53 (1983).","journal-title":"Proc. Nutr. Soc."},{"key":"e_1_3_1_7_2","unstructured":"A. Sommer in Elevated Dosages of Vitamins: Benefits and Hazards P. Walter G. Brubacher H. St\u00e4helin Eds. (Hans Huber Toronto Canada 1989) pp. 37\u201341."},{"key":"e_1_3_1_8_2","doi-asserted-by":"crossref","first-page":"1071","DOI":"10.1046\/j.1365-313X.1997.11051071.x","volume":"11","author":"Burkhardt P.","year":"1997","unstructured":"Burkhardt P., et al., Plant J. 11, 1071 (1997).","journal-title":"Plant J."},{"key":"e_1_3_1_9_2","unstructured":"Three vectors\u2014pUC18 pPZP100 and pBin19 (15\u201317)\u2014were digested with Eco RI and Hind III and a synthetic linker flanking by meganuclease I\u2013Sce I including Kpn I Not I and Sma I (5\u2032-AATTCATTACCCTGTTATCCCTACCCGGGCGGCCGCGGTACCATTACCCTGTTATCCCTAA-3\u2032) and (5\u2032-AGCTTTAGGGATAACAGGGTAATGGTACCGCGGCCGCCCGGGTAGGGATAACGGGTAATG-3\u2032) were introduced forming pUC18M pPZP100M and pBin19M respectively. An intermediate vector was made by insertion of the crtI expression cassette excised from Hind III\/Eco RI\u2013digested pUCET4 originally derived from pYPIET4 (11) into pBluescriptKS with Hind III\/Eco RI digestion followed by insertion of psy expression cassette from Sac II\u2013blunted\/Kpn I\u2013digested pGt1psyH (7) into the Kpn I\/Xho I\u2013blunted previous vector. Finally crtI and psy expression cassettes were isolated with Kpn I\/Not I digestion and inserted into Kpn I\/Not I\u2013digested pUC18M and designated as pBaal3. pBin19hpc was made by insertion of a Kpn I fragment originally from pCIB900 (18) containing aphIV selectable marker gene into pBaal3 followed by digestion of the I-Sce I fragment of the resulting plasmid and insertion into I-Sce I\u2013digested pBin19M. pZPsC was obtained by insertion of the I- Sce I fragment of pBaal3 bearing the psy and crtI genes into I-Sce I\u2013digested pPZP100M. pZLcyH was constructed by digestion of pGt1LcyH with I-Sce I and insertion of the resulting fragment carrying lcy and aphIV into I-Sce I\u2013restricted pPZP100M. The three vectors were separately electroporated into Agrobacterium tumefaciens LBA4404 (19) with corresponding antibiotic selection. Callus induction: Immature seeds of japonica rice cultivar TP 309 at milk stage were collected from greenhouse-grown plants surface-sterilized in 70% ethanol (v\/v) for 1 min incubated in 6% calcium hypochloride for 1 hour on a shaker and rinsed three to five times with sterile distilled water. Immature embryos were then isolated from the sterilized seeds and cultured onto NB medium [N6 salts and B5 vitamins supplemented with 30 g\/l maltose 500 mg\/l proline 300 mg\/l casein hydrolate 500 mg\/l glutamine and 2 mg\/l 2 4-D (pH 5.8)]. After 4 to 5 days the coleoptiles were removed and the swelled scutella were subcultured onto fresh NB medium for 3 to 5 days until inoculation of Agrobacterium. Transformation: 1-week-old precultured immature embryos were immersed in Agrobacterium tumefaciens LBA 4404 cell suspension as described (20). For co-transformation LBA4404\/pZPsC [optical density at 600 nm (OD 600 ) = 2.0] mixed with an equal volume of LBA4404\/pZLcyH (OD 600 = 1.0) was used for inoculation after acetonsyrigone induction. The inoculated precultured embryos were co-cultivated onto NB medium supplemented with 200 mM acetonsyringone for 3 days subcultured on recovery medium (NB with 250 mg\/l cefotaxime) for 1 week and then transferred onto NB selection medium in the presence of 30 mg\/l hygromycin and 250 mg\/l cefotaxime for 4 to 6 weeks. Transgenic plants were regenerated from recovered resistant calli on NB medium supplemented with 0.5 mg\/l NAA and 3 mg\/l BAP in 4 weeks rooted and transferred into the greenhouse."},{"key":"e_1_3_1_10_2","doi-asserted-by":"crossref","first-page":"781","DOI":"10.1046\/j.1365-313X.1996.10050781.x","volume":"10","author":"Schledz M.","year":"1996","unstructured":"Schledz M., et al., Plant J. 10, 781 (1996).","journal-title":"Plant J."},{"key":"e_1_3_1_11_2","doi-asserted-by":"crossref","first-page":"942","DOI":"10.1111\/j.1432-1033.1997.00942.x","volume":"247","author":"Bonk M.","year":"1997","unstructured":"Bonk M., et al., Eur. J. Biochem. 247, 942 (1997).","journal-title":"Eur. J. Biochem."},{"key":"e_1_3_1_12_2","doi-asserted-by":"crossref","first-page":"833","DOI":"10.1046\/j.1365-313X.1993.04050833.x","volume":"4","author":"Misawa N.","year":"1993","unstructured":"Misawa N., et al., Plant J. 4, 833 (1993).","journal-title":"Plant J."},{"key":"e_1_3_1_13_2","first-page":"1398","volume":"112","author":"Al-Babili S.","year":"1996","unstructured":"Al-Babili S., Hobeika E., Beyer P., Plant Physiol. 112, 1398 (1996).","journal-title":"Plant Physiol."},{"key":"e_1_3_1_14_2","unstructured":"Dehusked seeds were polished for 6 hours with emery paper on a shaker. The endosperm obtained was ground to a fine powder and 1 g was extracted repeatedly with acetone. Combined extracts were used to record the ultraviolet-visible spectrum allowing quantification using \u03b5 450nm 134 000 l \u20131 mol \u20131 cm \u20131 for \u03b2-carotene. The samples were dried and the residue quantitatively applied in 30 \u03bcl chloroform to HPLC for analysis using a photodiode array detector (Waters) and a C 30 reversed-phase column (YMC Europe GmbH) with the solvent system A [methanol: tert -butylmethyl ether (1:1 v\/v)] and system B [methanol: tert -butylmethyl ether:H 2 O (6:1.2:1.2 v\/v\/v)] using a gradient of 100% B to 43% B within 25 min then to 0% B within a further 75 min. Final conditions were maintained for an additional 10 min. Photometric quantifications were re-examined by HPLC using synthetic all- trans lycopene as an external standard."},{"key":"e_1_3_1_15_2","doi-asserted-by":"crossref","first-page":"57","DOI":"10.1006\/exer.1997.0309","volume":"65","author":"Landrum J. T.","year":"1997","unstructured":"Landrum J. T., et al., Exp. Eye Res. 65, 57 (1997).","journal-title":"Exp. Eye Res."},{"key":"e_1_3_1_16_2","doi-asserted-by":"crossref","first-page":"103","DOI":"10.1016\/0378-1119(85)90120-9","volume":"33","author":"Yanish-Perron C.","year":"1985","unstructured":"Yanish-Perron C., Vieira J., Messing J. C., Gene 33, 103 (1985).","journal-title":"Gene"},{"key":"e_1_3_1_17_2","doi-asserted-by":"publisher","DOI":"10.1007\/BF00014672"},{"key":"e_1_3_1_18_2","doi-asserted-by":"publisher","DOI":"10.1093\/nar\/12.22.8711"},{"key":"e_1_3_1_19_2","first-page":"171","volume":"14","author":"Wuenn J.","year":"1996","unstructured":"Wuenn J., et al., Bio\/Technology 14, 171 (1996).","journal-title":"Bio\/Technology"},{"key":"e_1_3_1_20_2","doi-asserted-by":"crossref","first-page":"179","DOI":"10.1038\/303179a0","volume":"303","author":"Hoekema A.","year":"1983","unstructured":"Hoekema A., Hirsch P. R., Hooykaas P. J. J., Schilperoort R. A., Nature 303, 179 (1983).","journal-title":"Nature"},{"key":"e_1_3_1_21_2","doi-asserted-by":"crossref","first-page":"87","DOI":"10.1016\/S0168-9452(97)00211-2","volume":"130","author":"Uze M.","year":"1997","unstructured":"Uze M., et al., Plant Sci. 130, 87 (1997).","journal-title":"Plant Sci."},{"key":"e_1_3_1_22_2","unstructured":"The crtI gene fused to the transit peptide sequence was kindly provided by N. Misawa (Kirin Co. Ltd. Japan). We thank W. Dong and P. Burkhardt for their valuable contributions S. Klarer K. Konja and U. Schneider-Ziebert for skillful technical assistance and R. Cassada for correcting the English version of the manuscript. Supported by the Rockefeller Foundation (1993\u20131996) the European Community Biotech Program (FAIR CT96 1996\u20131999) (P.B.) the Swiss Federal Office for Education and Science (I.P.) and by the Swiss Federal Institute of Technology (1993\u20131996)."}],"container-title":["Science"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.science.org\/doi\/pdf\/10.1126\/science.287.5451.303","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2024,1,13]],"date-time":"2024-01-13T05:06:38Z","timestamp":1705122398000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.science.org\/doi\/10.1126\/science.287.5451.303"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2000,1,14]]},"references-count":21,"journal-issue":{"issue":"5451","published-print":{"date-parts":[[2000,1,14]]}},"alternative-id":["10.1126\/science.287.5451.303"],"URL":"https:\/\/doi.org\/10.1126\/science.287.5451.303","relation":{},"ISSN":["0036-8075","1095-9203"],"issn-type":[{"value":"0036-8075","type":"print"},{"value":"1095-9203","type":"electronic"}],"subject":[],"published":{"date-parts":[[2000,1,14]]}}}