JOURNAL BROWSE
Search
Advanced SearchSearch Tips
Development of high tryptophan GM rice and its transcriptome analysis
facebook(new window)  Pirnt(new window) E-mail(new window) Excel Download
  • Journal title : Journal of Plant Biotechnology
  • Volume 42, Issue 3,  2015, pp.186-195
  • Publisher : The Korean Society of Plant Biotechnology
  • DOI : 10.5010/JPB.2015.42.3.186
 Title & Authors
Development of high tryptophan GM rice and its transcriptome analysis
Jung, Yu Jin; Nogoy, Franz Marielle; Cho, Yong-Gu; Kang, Kwon Kyoo;
  PDF(new window)
 Abstract
Anthranilate synthase (AS) is a key enzyme in the biosynthesis of tryptophan (Trp), which is the precursor of bioactive metabolites like indole-3-acetic acid and other indole alkaloids. Alpha anthranilate synthase 2 (OsASA2) plays a critical role in the feedback inhibition of tryptophan biosynthesis. In this study, two vectors with single (F124V) and double (S126F/L530D) point mutations of the OsASA2 gene for feedback-insensitive subunit of rice anthranilate synthase were constructed and transformed into wildtype Dongjinbyeo by Agrobacterium-mediated transformation. Transgenic single and double mutant lines were selected as a single copy using TaqMan PCR utilized nos gene probe. To select intergenic lines, the flanking sequence of RB or LB was digested with a BfaI enzyme. Four intergenic lines were selected using a flanking sequence tagged (FST) analysis. Expression in rice (Oryza sativa L.) of the transgenes resulted in the accumulation of tryptophan (Trp), indole-3-acetonitrile (IAN), and indole-3-acetic acid (IAA) in leaves and tryptophan content as a free amino acid in seeds also increased up to 30 times relative to the wildtype. Two homozygous event lines, S-TG1 and D-TG1, were selected for characterization of agronomic traits and metabolite profiling of seeds. Differentially expressed genes (DEGs), related to ion transfer and nutrient supply, were upregulated and DEGs related to co-enzymes that work as functional genes were down regulated. These results suggest that two homozygous event lines may prove effective for the breeding of crops with an increased level of free tryptophan content.
 Keywords
Anthranilate synthase;OsASA2 gene;Transgenic event rice;Tryptophan;
 Language
Korean
 Cited by
 References
1.
Bartel B (1997) Auxin biosynthesis. Annu. Rev. Plant Physiol. Plant Mol. Biol. 48:51-66 crossref(new window)

2.
Bender J, Fink GR (1998) A myb homologue, ATR1, activates tryptophan gene expression in Arabidopsis. Proceedings of the National Academy of Sciences, USA 95:5655-5660 crossref(new window)

3.
Bohlmann J, DeLuca V, Eilert U, Martin W (1995) Purification and cDNA cloning of anthranilate synthase from Ruta graveolens: modes of expression and properties of native and recombinant enzymes. Plant J 7:491-501 crossref(new window)

4.
Hattori A, Migitaka H, Iigo M, Itoh M, Yamamoto K, Ohtanikaneko R, Hara M, Suzuki T, Reiter RJ (1995) Identification of melatonin in plants and its effects on plasma melatonin levels and binding to melatonin receptors in vertebrates. Biochemistry and Molecular Biology International 35:627-634

5.
Huber W, von Heydebreck A, Sultmann H, Poustka A, Vingron M (2002) Variance stabilization applied to microarray data calibration and to the quantification of differential expression. Bioinformatics 18 (Suppl 1): S96-S104 crossref(new window)

6.
Ishihara A, Matsuda F, Miyagawa H, Wakasa K (2007) Metabolomics for metabolically manipulated plants: effects of tryptophan overproduction. Metabolomics 3:319-334 crossref(new window)

7.
Jung YJ, Lee MC, Kang KK (2011) A transcription factor "OsNAC075" is essential for salt resistance in rice (Oryza sativa L.). J Plant Biotechnol 38:94-104 crossref(new window)

8.
Jung YJ, Nou IS, Kang KK (2014) Overexpression of Oshsp16.9 gene encoding small heat shock protein enhances tolerance to abiotic stresses in rice. Plant Breed. Biotech. 2(4):370-379 crossref(new window)

9.
Keay S, Seillier-Moiseiwitsch F, Zhang CO, Chai TC, Zhang J (2003) Changes in human bladder epithelial cell gene expression associated with interstitial cystitis or antiproliferative factor treatment. Physiological Genomics 14:107-115 crossref(new window)

10.
Kim DS, Lee IS, Jang CS, Kang SY, Seo YW (2005) Characterization of the altered anthranilate synthase in 5-methyltryptophan- resistant rice mutants. Plant Cell Rep 24:357-365 crossref(new window)

11.
Lee HJ, Abdula SE, Jee MG, Jang DW, Cho YG (2011) High-efficiency and Rapid Agrobacterium-mediated genetic transformation method using germinating rice seeds. J Plant Biotechnol 38:251-257 crossref(new window)

12.
Lee HY, Kameya T (1991) Selection and characterization of a rice mutant resistant to 5-methyltryptophan. Theor Appl Genet 82:405-408 crossref(new window)

13.
Livak KJ, Schmittgen TD (2001) Analysis of relative gene expression data using real-time quantitative PCR and the $2^{-{\Delta}{\Delta}Ct}$ method. Methods 25:402-408 crossref(new window)

14.
Murch SJ, KrishnaRaj S, Saxena PK (2000) Tryptophan is a precursor for melatonin and serotonin biosynthesis in in vitro regenerated St John's wort (Hypericum perforatum L. cv. Anthos) plants. Plant Cell Reporter 19:698-704 crossref(new window)

15.
Radwanski ER, Last RL (1995) Tryptophan biosynthesis and metabolism: biochemical and molecular genetics. Plant Cell 7:921-934 crossref(new window)

16.
Romero RM, Roberts MF (1996) Anthranilate synthase from Ailanthus altissima cell suspension cultures. Phytochemistry 41:395-402 crossref(new window)

17.
Thole V, Alves SC, Worland B, Bevan MW, Vain P (2009) A protocol for efficiently retrieving and characterizing flanking sequence tags (FSTs) in Brachypodium distachyon T-DNA insertional mutants. Nature Protocols 4:650-661 crossref(new window)

18.
Tozawa Y, Hasegawa H, Terakawa T, Wakasa K (2001) Characterization of rice anthranilate synthase a-subunit genes OASA1 and OASA2. Tryptophan accumulation in transgenic rice expressing a feedback-insensitive mutant of OASA1. Plant Physiology 126:1493-1506 crossref(new window)

19.
Ueno M, Shibata H, Kihara J, Honda Y, Arase S (2003) Increased tryptophan decarboxylase and monoamine oxidase activities induce Sekiguchi lesion formation in rice infected with Magnaporthe grisea. The Plant Journal 36: 215-228 crossref(new window)

20.
Wakasa K, Hasegawa H, Nemoto H, Matsuda F, Miyazawa H, Tozawa Y, Morino K, Komatsu A, Yamada T, Terakawa T, Miyagawa (2006) High-level tryptophan accumulation in seeds of transgenic rice and its limited effects on agronomic traits and seed metabolite profile. J Exp Bot 57:3069-3078 crossref(new window)

21.
Wakasa K, Ishihara A (2009) Metabolic engineering of the tryptophan and phenylalanine biosynthetic pathways in rice. Plant Biotechnol 26:523-533 crossref(new window)

22.
Wakasa K, Widholm JM (1987) A 5-methyltryptophan resistant rice mutant, MTR1, selected in tissue-culture. Theor Appl Genet 74:49-54 crossref(new window)