Welcome to International Network for Natural Sciences | INNSpub

Molecular cloning and expression profiling of a chalcone synthase gene from Ginkgo biloba, Scutellaria viscidula Bunge and Tree peony (Paeonia suffruticosa)

Review Paper | October 1, 2013

| Download 3

Manaf Almatar

Key Words:

Int. J. Biosci.3( 10), 187-202, October 2013

DOI: http://dx.doi.org/10.12692/ijb/3.10.187-202


IJB 2013 [Generate Certificate]


Chalcone synthase (CHS, EC is a key enzyme of the flavonoid/isoflavonoid biosynthesis pathway. In this review, the characterization and expression profile of CHS was investigated from Ginkgo biloba, Scutellaria viscidula, and Tree peony (Paeonia suffruticosa) because of the CHS role in protecting the plants from harmful effects by synthesizing flavonoid compounds. CHS was isolated and characterized from the same species which was described previously. The cDNA sequence of the CHS gene in Ginkgo biloba was 1608 bp with poly (A) tailing, from Scutellaria viscidula1649 bp, and from Tree peony (Paeonia suffruticosa) 1475 bp. cDNA sequence of the CHS gene in G. biloba contained a 1173 bp open reading frame (ORF) encoding a 391 amino acid protein, from Scutellaria viscidula 1170 bp encoding a 390 amino acid protein, and from Tree peony (Paeonia suffruticosa) 1185bp encoding a 394 amino acid protein. Sequence alignment and phylogenetic analysis showed that GbCHS, SvCHS and Ps-CHS1 shared high homology from other plants. Active site of the CoA binding, coumaroyl pocket and cyclization pocket in CHS protein of Medicago sativa were also found in GbCHS, and SvCHS. Likewise, Ps-CHS1 has all the conserved active sites for the CHS function as well as the family signature. The homology-based structural modeling showed that GbCHS most closely resembled of M.sativa (MsCHS2). Real-time (PCR) analysis indicated that GbCHS and Ps-CHS1 had varying expression in different tissues, while expression of SvCHS was induced by MeJA and was the greatest at 12 h. To our knowledge, this is the first report which explains the molecular cloning and expression profiling of a chalcone synthase gene from those species mentioned above.


Copyright © 2013
By Authors and International Network for
Natural Sciences (INNSPUB)
This article is published under the terms of the Creative
Commons Attribution Liscense 4.0

Molecular cloning and expression profiling of a chalcone synthase gene from Ginkgo biloba, Scutellaria viscidula Bunge and Tree peony (Paeonia suffruticosa)

Ananieva K, Ananiev ED. 1997. Comparative study of the effects of methyl jasmonate and abscisic acid on rna and protein synthesis in excised cotyledons of Cucurbita Pepo L. (ZUCCHINI). Bulg. J. Plant physiol 23, 80–90.

Arnold K, Bordoli L, Kopp J, Schwede T. 2006. The SWISS-MODEL workspace: A web-based environment for protein structure homology modelling. Bioinformatics 22, 195-201. http://dx.doi.org/ 10.1093/bioinformatics/bti770

Austin MB, Noel JP. 2003. The chalcone synthase superfamily of type III polyketide synthases. Natural Product Reports 20, 79-110. http://www.ncbi.nlm.nih.gov/pubmed/12636085

Belhadj A, Telef N, Saigne C, Cluzet S, Barrieu F, Hamdi S, Mérillon JM. 2008. Effect of methyl jasmonate in combination with carbohydrates on gene expression of PR proteins, stilbene and anthocyanin accumulation in grapevine cell cultures. Plant Physiology and Biochemistry 46, 493-499. http://dx.doi.org/10.1016/j.plaphy.2007.12.001

Bilgin DD, DeLucia EH, Clough SJ. 2009. A robust plant RNA isolation method suitable for Affymetrix GeneChip analysis and quantitative real-time RT-PCR. Nature protocols 4, 333-340. http://dx.doi.org/ 10.1038/nprot.2008.249

Burland TG. 1999. DNASTAR’s Lasergene Sequence Analysis Software. In: Misener S, Krawetz SA, Editors. Bioinformatics Methods and Protocols. New Jersy: Humana Press 71-91.

Bustin SA. 2000. Absolute quantification of mRNA using real-time reverse transcription polymerase chain reaction assays. Journal of Molecular Endocrinology 25, 169-193. http://dx.doi.org/ 10.1677/jme.0.0250169

Cameron M, Williams H. 2007. Comparing Compressed  Sequences  for  Faster  Nucleotide  blast Searches. Ieee/acm trans comput biol Bioinformatics 4,349-364. http://doi.ieeecomputersociety.org/10.1109/TCBB.2007.1029

Cao S, Cai Y, Yang Z, Zheng Y. 2012.  MeJA induces chilling tolerance in loquat fruit by regulating proline and γ-aminobutyric acid contents. Food Chemistry 133, 1466-1470. http://dx.doi.org/10.1016/j.foodchem.2012.02.035

Chang S, Puryear J, Cairney J. 1993. A simple and efficient method for isolating RNA from pine trees. Plant Molecular Biology Reporter 11, 113-116. http://dx.doi.org/ 10.1007/BF02670468

Claudot AC, Ernst D, Sandermann HJR, Drouet A. 1999. Cloning and characterization of two members of the chalcone synthase gene family from walnut. Plant Physiology and Biochemistry 37, 721-730. http://dx.doi.org/10.1016/S0981-9428(00)86685-1

Creelman RA, Tierney ML, Mullet JE. 1992. Jasmonic acid/methyl jasmonate accumulate in wounded soybean hypocotyls and modulate wound gene expression. Proceedings of the National Academy of Sciences of the United States of America 89, 4938-4941.

Deepak SA, Kottapalli KR, Rakwal R, Oros G, Rangappa KS, Iwahashi H, Masuo Y, Agrawal GK. 2007. Real-time PCR: Revolutionizing detection and expression analysis of genes. Current Genomics 8, 234-251.

Felsenstein J. 1985. Confidence Limits on Phylogenies: An Approach Using the Bootstrap. Evolution 39, 783-791.

Ferrer JL, Jez JM, Bowman ME, Dixon RA, Noel JP. 1999. Structure of chalcone synthase and the molecular basis of plant polyketide biosynthesis. Nature Structural Biology 6, 775-784. http://dx.doi.org/ 10.1038/11553

Fliegmann J, Schröder G, Schanz S, Britsch L, Schröder J. 1992. Molecular analysis of chalcone and dihydropinosylvin synthase from Scots pine (Pinus sylvestris), and differential regulation of these and related enzyme activities in stressed plants. Plant Molecular Biology 18, 489-503. http://dx.doi.org/ 10.1007/BF00040665

Franken P, Niesbach-Klösgen U, Weydemann U, Maréchal-Drouard L, Saedler H, Wienand U. 1991. The duplicated chalcone synthase genes C2 and Whp (white pollen) of Zea mays are independently regulated; Evidence for translational control of Whp expression by the anthocyanin intensifying gene in. EMBO Journal 10, 2605-2612.

Frohman MA, Dush MK, Martin GR. 1988. Rapid production of full-length cDNAs from rare transcripts: amplification using a single gene-specific oligonucleotide primer. Proceedings of the National Academy of Sciences 85, 8998-9002.

Goto-Yamamoto N, Wan GH, Masaki K, Kobayashic S. 2002. Structure and transcription of three chalcone synthase genes of grapevine (Vitis vinifera). Plant Science 162, 867-872. http://dx.doi.org/10.1016/S0168-9452(02)00042-0

Helariutta Y, Elomaa P, Kotilainen M, Griesbach RJ, Schröder J, Teeri TH. 1995. Chalcone synthase-like genes active during corolla development are differentially expressed and encode enzymes with different catalytic properties in Gerbera hybrida (Asteraceae). Plant Molecular Biology 28, 47-60. htt://dx.doi.org/10.1007/BF00042037

Hillmann A, Dunne E, Kenny D. 2009. cDNA amplification by SMART-PCR and suppression subtractive hybridization (SSH)-PCR, P. Bugert, Editor 496, 223-243. http://dx.doi.org/ 10.1007/978-1-59745-553-4_15

Hua YG, Bin L, Wei GJ, Zhong LJ, Qiang ZX, Qi Z, Yi-Ping T, Zhen-Sheng L. 2004. Cloning and expression of two chalcone synthase and a flavonoid 3′5′-hydroxylase 3′-end cDNAs from developing seeds of blue-grained wheat involved in anthocyanin biosynthetic pathway. Acta Botanica Sinica 46, 588-594.

Huang B, Yi B, Duan Y, Sun L, Yu X, Guo J, Chen W. 2008. Characterization and expression profiling of tyrosine aminotransferase gene from Salvia miltiorrhiza (Dan-shen) in rosmarinic acid biosynthesis pathway. Molecular Biology Reports 35, 601-612. http://dx.doi.org/ 10.1007/s11033-007-9130-2

Huang J, Gu M, Lai Z, Fan B, Shi K, Zhou YH, Yu JQ, Chen Z. 2010.Functional analysis of the Arabidopsis PAL gene family in plant growth, development, and response to environmental stress. Plant Physiology 153, 1526-538. http://dx.doi.org/10.1104/pp.110.157370

Jaakola L, Pirttilä AM, Halonen M, Hohtola A. 2001. Isolation of high quality RNA from bilberry (Vaccinium myrtillus L.) fruit. Applied Biochemistry and Biotechnology – Part B Molecular Biotechnology 19, 201-203. http://dx.doi.org/ 10.1385/MB:19:2:201

Jez JM,Noel JP. 2000. Mechanism of Chalcone Synthase: pk a of the catalytic cysteine and the role of the conserved histidine in a plant polyketide synthase. Journal of Biological Chemistry 275, 39640-39646. http://dx.doi.org/ 10.1074/jbc.M008569200

Jiang C, Schommer CK, Kim SY, Suh DY. 2006. Cloning  and  characterization  of  chalcone  synthase from the moss, Physcomitrella patens. Phytochemistry 67, 2531-2540. http://dx.doi.org/10.1016/j.phytochem.2006.09.030

Jiang M, Cao SA. 2007. Chalcone Synthase Gene. Chinese Journal of Cell Biology 29, 525-529.

Kalpesh A. 2012. DNA sequence analysis by orf finder & genomatix tool: Bioinformatics analysis of some tree species of Leguminosae family.IEEE Computer Society Washington, DC, USA 922-926. http://dx.doi.org/ 10.1109/BIBMW.2012.6470265

Kashkush K, Feldman M, Levy AA. 2003. Transcriptional activation of retrotransposons alters the expression of adjacent genes in wheat. Nature Genetics 33, 102-106. http://dx.doi.org/ 10.1038/ng1063

Kim SH, Mizuno K, Fujimura T. 2002. Regulated expression of ADPglucose pyrophosphorylase and chalcone synthase during root development in sweet potato. Plant Growth Regulation 38, 173-179.

Kovács Gy, Kuzovkina IN, Szoke É, Kursinszki L. 2004. HPLC determination of flavonoids in hairy-root cultures of Scutellaria baicalensis Georgi. Chromatographia 60, 81-85. http://dx.doi.org/ 10.1365/s10337-004-0266-0

Kumar S, Tamura K, Jakobsen IB, Nei M. 2001. MEGA2: molecular evolutionary genetics analysis software. Bioinformatics 17,1244-1245. http://dx.doi.org/ 10.1093/bioinformatics/17.12.1244

Kumar S, Tamura K, Nei M. 2004. MEGA3: Integrated software for Molecular Evolutionary Genetics Analysis and sequence alignment. Briefings in Bioinformatics 5, 150-163. http://dx.doi.org/ 10.1093/bib/5.2.150

Kvacek J, Falcon-Lang HJ, Dasková J. 2005. A new Late Cretaceous ginkgoalean reproductive structure Nehvizdyella gen. nov. from the Czech Republic and its whole-plant reconstruction. American Journal of Botany 92,1958-1969. http://dx.doi.org/ 3732/ajb.92.12.1958

Lanz T, Tropf S, Marner FJ, Schröder J, Schröder G. 1991. The role of cysteines in polyketide synthases: Site-directed mutagenesis of resveratrol and chalcone synthases, two key enzymes in different plant-specific pathways. Journal of Biological Chemistry 266, 9971-9976.

Lei W, Sun M, Luo KM, Shui XR, Sun YM, Tang H. 2009. Compute simulation to characterize structure and function of chalcone synthase from Scutellaria baicalensis Georgi. Molekuliarnaia biologiia 43,1082-1087.

http://dx.doi.org/ 10.1134/S0026893309060144

Lei W, Tang SH, Luo KM, Sun M. 2010. Molecular cloning and expression profiling of a chalcone synthase gene from hairy root cultures of Scutellaria viscidula Bunge. Genetics and Molecular Biology 33, 285-291.

Liao Z, Chen M, Guo L, Gong Y, Tang F, Sun X, Tang K. 2004.Rapid isolation of high-quality total RNA from taxus and ginkgo. Preparative Biochemistry and Biotechnology 34, 209-214.

Liew CF,Goh CJ, Loh CS, Lim SH. 1998. Cloning and characterization of full-length cDNA clones encoding chalcone synthase from the orchid Bromheadia finlaysoniana. Plant Physiology and Biochemistry 36, 647-656. http://dx.doi.org/10.1016/S0981-9428(98)80013-2

Li KB. 2003. ClustalW-MPI: ClustalW analysis using distributed and parallel computing. Bioinformatics 19, 1585-1586. http://dx.doi.org/ 10.1093/bioinformatics/btg192

Luscombe  NM,  Greenbaum  D,  Gerstein  M. 2001. What is bioinformatics? An introduction and overview. Yearbook of Medical Informatics 83-99.

Mahadevan S, Park Y. 2008. Multifaceted Therapeutic Benefits of Ginkgo biloba L. Chemistry, Efficacy, Safety, and Uses. Journal of Food Science 73, 14-19. http://dx.doi.org/ 10.1111/j.1750-3841.2007.00597.x

Makoi JH,Bambara S, Ndakidemi PA. 2010. Rhizosphere phosphatase enzyme activities and secondary metabolites in plants as affected by the supply of Rhizobium, lime and molybdenum in Phaseolus vulgaris L. Australian Journal of Crop Science 4, 590-597.

Martens S, Mithöfer A. 2006. Erratum: Flavones and flavone synthases (Phytochemistry (2005) 66 (2399-2407) DOI:10.1016/j.phytochem.2005.07.013). Phytochemistry 66, 2399-2407.

Moreira X, Sampedro L, Zas R. 2009. Defensive responses of Pinus pinaster seedlings to exogenous application of methyl jasmonate: Concentration effect and systemic response. Environmental and Experimental Botany 67, 94-100. http://dx.doi.org/10.1016/j.envexpbot.2009.05.015

Mornya PMP,Cheng FY, Li HY. 2011. Chronological changes in plant hormone and sugar contents in cv. Ao-Shuang autumn flowering tree peony. Horticultural Science 38, 104-112.

Nakatsuka A, Izumi Y, Yamagishi M. 2003. Spatial and temporal expression of chalcone synthase and dihydroflavonol 4-reductase genes in the Asiatic hybrid lily. Plant Science 165, 759-767. http://dx.doi.org/10.1016/S0168-9452(03)00254-1

Napoli CA, Fahy D, Wang HY, Taylor LP. 1999. White anther: A Petunia Mutant That Abolishes Pollen Flavonol Accumulation, Induces Male Sterility, and Is Complemented by a Chalcone Synthase Transgene. Plant Physiology 120, 615-622. http://dx.doi.org/10.1104/pp.120.2.615

O’Neill SD, Tong Y, Spörlein B, Forkmann G, Yoder JI. 1990. Molecular genetic analysis of chalcone synthase in Lycopersicon esculentum and an anthocyanin-deficient mutant. Molecular and General Genetics 224, 279-288. http://dx.doi.org/ 10.1007/BF00271562

Pang Y, Shen G, Wu W, Liu X, Lin J, Tan F, Sun X, Tang K. 2005. Characterization and expression of chalcone synthase gene from Ginkgo biloba. Plant Science 168, 1525-1531.

Pang Y, Shen GA, Liu C, Liu X, Tan F, Sun X, Tang K. 2004. Molecular cloning and sequence analysis of a novel chalcone synthase cDNA from Ginkgo biloba. DNA Sequence – Journal of DNA Sequencing and Mapping 15, 283-290.

Rajeevan MS, Ranamukhaarachchi DG, Vernon SD, Unger ER. 2001.  Use of  real-time quantitative pcr to validate the results of cDNA array and differential display PCR technologies. Methods 25, 443-451. http://dx.doi.org/10.1006/meth.2001.1266

Saitou N, Nei M. 1987. The neighbor-joining method: a new method for reconstructing phylogenetic trees. Molecular biology and evolution 4, 406-425.

Sánchez-Sampedro MA, Fernández-Tárrago J, Corchete P. 2005. Yeast extract and methyl jasmonate-induced silymarin production in cell cultures of Silybum marianum (L.) Gaertn. Journal of Biotechnology 119,60-69. http://dx.doi.org/10.1016/j.jbiotec.2005.06.012

Saslowsky DE,  Dana  CD,  Winkel-Shirley B. 2000. An allelic series for the chalcone synthase locus in Arabidopsis. Gene 255, 127-138. http://dx.doi.org/10.1016/S0378-1119(00)00304-8

SatiS C, Joshi S. 2011. Antibacterial activities of Ginkgo biloba L. leaf extracts. The Scientific World Journal 11, 2237-2242.

Schröder J, Raiber S, Berger T, Schmidt A, Schmidt J, Soares-Sello AM, Schwede T, Kopp J, Guex N, Peitsch MC. 2003. SWISS-MODEL: An automated protein homology-modeling server. Nucleic Acids Research 31, 3381-3385. http://dx.doi.org/ 10.1093/nar/gkg520

Schröder J, Raiber S, Berger T, Schmidt A, Schmidt J, Soares-Sello AM, Bardshiri E, Strack D, Simpson TJ, Veit M, Schröder G. 1998. Plant polyketide synthases: A chalcone synthase-type enzyme which performs a condensation reaction with methylmalonyl-Coa in the biosynthesis of C-methylated chalcones. Biochemistry 37, 8417-8425. http://dx.doi.org/ 10.1021/bi980204g

Ståhlberg A, Kubista M, Pfaffl M. 2004. Comparison of Reverse Transcriptases in Gene Expression Analysis. Clinical Chemistry 50,1678-1680. http://dx.doi.org/ 10.1373/clinchem.2004.035469

Takeuchi A, Matsumoto S, Hayatsu M. 1994. Chalcone Synthase from Camellia sinensis: Isolation of the cDNAs and the Organ-Specific and Sugar-Responsive Expression of the Genes. Plant and Cell Physiology 35:1011-1118.

Wang LS, Shiraishi A,Hashimoto F, Aoki N,Shimizu K, Sakata Y. 2001. Analysis of petal anthocyanins to investigate flower coloration of Zhongyuan (Chinese) and Daikon Island (Japanese) tree peony cultivars. Journal of Plant Research 114, 33-43. http://dx.doi.org/ 10.1007/PL00013966

Wang SY, Bowman L, Ding M. 2008. Methyl jasmonate enhances antioxidant activity and flavonoid content in blackberries (Rubus sp.) and promotes antiproliferation of human cancer cells. Food Chemistry 107, 1261-1269. http://dx.doi.org/10.1016/j.foodchem.2007.09.065

ang X, Cheng C, Sun Q, Li F, Liu J, Zheng C. 2005. Isolation and purification of four flavonoid constituents from the flowers of Paeonia suffruticosa by high-speed counter-current chromatography. Journal of Chromatography A 1075,127-131. http://dx.doi.org/10.1016/j.chroma.2005.04.017

Wilkins MR, Gasteiger E, Bairoch A, Sanchez JC, Williams KL, Appel RD, Bairoch A. 1999. Protein identification and analysis tools in the ExPASy server. Methods in molecular biology (Clifton, N.J.) 112, 531-52. http://dx.doi.org/ 10.1385/1-59259-890-0:571

Winkel-ShirleyB. 2002.Biosynthesis of flavonoids and effects of stress. Current Opinion in Plant Biology 5, 218-223. http://dx.doi.org/10.1016/S1369-5266(02)00256-X

Wu Q, Sun C, Chen S. 2012. Identification and expression analysis of a 3-hydroxy-3-methylglutaryl coenzyme A reductase gene from American ginseng. Plant OMICS 5, 414-420.

Xu F, Cai R, Cheng SY, Du H, Wang Y, Cheng SH. 2008. Molecular cloning, characterization and expression of phenylalanine ammonia-lyase gene from Ginkgo biloba. African Journal of Biotechnology 7, 721-729.

Xu F, Li L, Zhang W, Cheng H, Sun N, Cheng S, Wang Y. 2012. Isolation, characterization, and function analysis of a flavonol synthase gene from Ginkgo biloba. Molecular Biology Reports 39, 2285-2296. http://dx.doi.org/ 10.1007/s11033-011-0978-9

Yang C, Xu YR, Yao WX. 2002. Extraction of pharmaceutical components from Ginkgo biloba leaves using supercritical carbon dioxide. Journal of Agricultural and Food Chemistry 50, 846-849. http://pubs.acs.org/doi/abs/10.1021/jf010945f

Yang S, Terachi T,Yamagishi H. 2008. Inhibition of chalcone synthase expression in anthers of Raphanus sativus with ogura male sterile cytoplasm. Annals of Botany 102, 483-489. http://dx.doi.org/ 10.1021/jf010945f

Ye F, Xui L, Yi J, Zhang W, Zhang DY. 2002. Anticancer activity of Scutellaria baicalensis and its potential mechanism. Journal of Alternative and Complementary Medicine 8, 567-572. http://dx.doi.org/ 10.1089/107555302320825075.

Zheljazkov VD, Cantrell CL, Ebelhar Mw,Coker C,Evans WB. 2007. Quality Assessment and Yield of Baikal Skullcap (Scutellaria baicalensis) Grown at Multiple Locations. HortScience 42, 1183-1187.

Zhou L, Wang Y, Peng Z. 2011. Molecular characterization and expression analysis of chalcone synthase  gene  during  flower  development  in  tree peony (paeonia suffruticosa). African Journal of Biotechnology 10, 1275-1284. http://dx.doi.org/ 10.5897/AJB10.599