Overview

Within the realm of synthetic biology, cells cease to be mere observers and act as dedicated craftsmen, assembling novel molecular machines under experimental direction.

The iGEM 2025 JLU-NBBMS registry includes 23 basic parts and 12 composite parts designed for the biosynthesis of ginsenoside Ro from glycerol and fatty acids. These parts exemplify our synthetic biology approach, emphasizing modular and standardized gene expression to achieve predictable outcomes across biological systems. These parts embody our collective effort and wisdom, enabling us to achieve success in the laboratory work of our project.

Basic parts

Number	Short Description	Function Description
BBa_25P0NBEQ	ADH1 promotor	The ADH1 promoter (pADH1) is a DNA sequence from the alcohol dehydrogenase 1 gene that drives strong, constitutive expression in yeast. It recruits transcription factors and RNA polymerase II to initiate transcription, making it widely used for protein overexpression and metabolic engineering.
BBa_K3126002	GPM1 promotor	The GPM1 promoter (pGPM1) provides binding sites for RNA polymerase II and a series of universal transcription factors, forming a transcription initiation complex to initiate the transcription process of the target gene.
BBa_K5058012	TEF1 promotor	The TEF1 promoter (pTEF1) forms a transcription initiation complex, initiating transcription of the target gene and generating substantial amounts of mRNA.
BBa_K3112025	ADH1 terminator	The ADH1 terminator is a DNA sequence from the alcohol dehydrogenase 1 gene that signals transcription termination in yeast. It ensures proper release of RNA polymerase II and stabilizes the mRNA, promoting efficient gene expression and preventing read-through into downstream sequences
BBa_K4121046	CYC1 terminator	CYC1 encodes cytochrome c, a key component of the mitochondrial electron transport chain. The CYC1 terminator ensures precise transcription termination, prevents transcription slippage, and promotes correct 3'-end processing, thereby enhancing mRNA stability and translational efficiency.
BBa_258302MX	(GGGGS)2 linker-GGC	The linker sequence represents a typical flexible linker, primarily composed of glycine (Gly) and serine (Ser). In fusion proteins, such linkers act as ‘spacers’: they effectively minimize steric or conformational interference between adjacent functional domains while preserving their independent folding and activity.
BBa_25FL7X0X	(GGGGS)2 linker-GGT	The linker sequence represents a typical flexible linker, primarily composed of glycine (Gly) and serine (Ser). In fusion proteins, such linkers act as ‘spacers’: they effectively minimize steric or conformational interference between adjacent functional domains while preserving their independent folding and activity.
BBa_K3819051	6×His Tag	The 6xHis tag facilitates protein purification and makes it convenient to extract target proteins from thousands of proteins in cells or cell lysates. Furthermore,the 6xHis tag has a small structure, so it is less likely to affect the function of the fusion protein, and a larger tag is more likely to affect the function of the fusion protein
BBa_251JGW0E	5×Leu Tag	The 5xLeu tag facilitates protein purification and makes it convenient to extract target proteins from thousands of proteins in cells or cell lysates. Furthermore,the 5xLeu tag has a small structure, so it is less likely to affect the function of the fusion protein, and a larger tag is more likely to affect the function of the fusion protein
BBa_25MXP281	Myc tag	The MYC tag is a synthetic short peptide sequence (EQKLISEEDL) widely employed in molecular biology. Its primary functions include facilitating Western blot analysis, immunoprecipitation, and immunofluorescence detection of proteins via anti-MYC antibodies.
BBa_25MWUYJC	Peroxisomal Targeting Signal 1 (PTS1)	PTS1 is a tripeptide sequence located at the C-terminus of proteins (typically SKL or its variants). Its primary function is to act as a ‘molecular postcode’, recognised by receptors on the peroxisomal membrane. This enables the specific targeting and transport of proteins bearing this sequence into the peroxisome lumen, where they perform the organelle's characteristic metabolic functions.
BBa_25HSNOC2	Acetoacetyl-CoA thiolase (Erg10)	ERG10 is the key initiating enzyme in the mevalonate (MVA) pathway of Saccharomyces cerevisiae, encoding acetyl-CoA acetyltransferase (AACT). Its core function is to catalyse the condensation of two molecules of acetyl-CoA to form acetoacetyl-CoA, which is the primary step in the synthesis of sterols (such as ergosterol) and terpenoid compounds, and is essential for maintaining cell membrane integrity.
BBa_25F0OIWH	Hydroxymethylglutaryl-CoA synthase (Erg13)	ERG13 is a key enzyme in the mevalonate (MVA) pathway of Saccharomyces cerevisiae, encoding hydroxymethylglutaroyl-CoA synthase . Its function is to catalyse the condensation of acetoacetyl-CoA with acetyl-CoA, yielding hydroxymethylglutaroyl-CoA (HMG-CoA). This represents an essential step in the synthesis of sterols (such as ergosterol) and terpenoid compounds, and is crucial for cellular membrane function.
BBa_25GQA97K	Hydroxymethylglutaryl-CoA reductase(HMG1)	HMG1 is the rate-limiting enzyme in the mevalonate pathway of Saccharomyces cerevisiae, encoding hydroxymethylglutaryl-coenzyme A reductase . Its core function is to catalyse the reduction of HMG-CoA to mevalonic acid, consuming NADPH in the process. This constitutes the first critical and irreversible step in the biosynthesis of sterols such as ergosterol, determining the rate of the entire metabolic pathway and being essential for maintaining cell membrane integrity.
BBa_25IC6WOS	Beta - amyrin synthase (β-AS)	It oxidizes 2,3-oxidosqualene to form β-amyrin, which is necessary for the synthesis of ginsenoside Ro's precursor oleanolic acid (OA).
BBa_25XK6L40	Cytochrome P450 monoxygenase (CYP716A12)	Together with cytochrome P450 reductase 1, it directly participates in the oxidation of β-amyrin to oleanolic acid, thereby providing the precursor for the biosynthesis of calenduloside E (CE).
BBa_25ZMN81X	Cytochrome P450 reductase 1(MtCPR)	Together with Cytochrome P450 monoxygenase, it directly participates in the oxidation of β-amyrin to oleanolic acid, thereby providing the precursor for the biosynthesis of calenduloside E (CE).
BBa_25H2OJ14	UGT73P40	UGT73P40 is a glycosyltransferase derived from the plant Panax notoginseng. When heterologously expressed in yeast, it catalyses the glycosylation of intermediate products (such as terpenoids), adding a sugar residue derived from uridine diphosphate glucose (UDP-glucose) to these molecules. This process is utilised in synthetic biology for the production of high-value glycoside compounds. In this project, the enzyme serves as one of the key catalysts for the generation of Ro.
BBa_25JN45NN	UGT73F3	UGT73F3 is a glycosyltransferase derived from the plant Panax notoginseng. When heterologously expressed in yeast, it catalyses the glycosylation of intermediate products (such as terpenoids), adding a sugar residue derived from uridine diphosphate glucose (UDP-glucose) to these molecules. This process is utilised in synthetic biology for the production of high-value glycoside compounds. In this project, the enzyme serves as one of the key catalysts for the generation of Ro.
BBa_250REJW3	Pn022859	Pn022859 is a glucuronosyltransferase derived from the plant Panax notoginseng. When heterologously expressed in yeast, it catalyses the glucuronidation of intermediates such as terpenoids—adding a uridine diphosphate glucuronic acid (UDP-GlcA) donor group to these molecules. This process is employed in synthetic biology for producing high-value glycoside compounds. Within this project, the enzyme serves as one of the key catalysts for generating Ro.
BBa_252VR4CL	Fructose-1,6-bisphosphatase(FBP-ase1)	FBP-ase1 is the key rate-limiting enzyme in the gluconeogenesis pathway of Saccharomyces cerevisiae. It catalyses the hydrolysis of fructose-1,6-bisphosphate (FBP) into fructose-6-phosphate and inorganic phosphate, an irreversible step constituting the decisive process by which cells synthesise glucose from non-sugar precursors. The activity of this enzyme is strongly inhibited by AMP and fructose-2,6-bisphosphate, ensuring the shutdown of the energy-consuming gluconeogenesis process during periods of sufficient energy availability.
BBa_25WFJC5W	Phosphoglucomutase(PGM2)	PGM2 encodes phosphoglucose isomerase in Saccharomyces cerevisiae, a pivotal enzyme in both glycolysis and gluconeogenesis. It catalyses the reversible conversion between glucose-1-phosphate and glucose-6-phosphate, a reaction that serves as the pivotal link connecting glycogen metabolism, glycolysis, and the pentose phosphate pathway. This reaction is crucial for the efficient utilisation of carbon sources and the maintenance of energy metabolism equilibrium.
BBa_25KBSR24	UTP glucose-1-phosphate uridylyltransferase (UGP1)	UGP1 is the key enzyme encoding UDP-glucose pyrophosphorylase in Saccharomyces cerevisiae. It catalyses the reversible reaction between glucose-1-phosphate and UTP to form UDP-glucose (UDP-Glc). UDP-Glc serves as a core precursor for glycogen, trehalose, and cell wall glucan synthesis. Consequently, UGP1 acts as a pivotal link between central carbon metabolism and cell polysaccharide synthesis, playing a crucial role in cellular energy storage, stress resistance, and structural integrity.

Composite parts

Number	Short Description	Function Description
BBa_25LZ47GL	ADH1 Promotor-6×His-linker-Erg10-2 linker-PTS1-ADH1 terminator	Since acetyl-CoA cannot freely diffuse across the peroxisomal membrane, this composite part enables the localization of ERG10 within peroxisomes, thereby allowing direct utilization of acetyl-CoA derived from peroxisomal fatty acid β-oxidation. ERG10 catalyzes the condensation of two acetyl-CoA molecules to form acetoacetyl-CoA, the precursor of HMG-CoA. In addition, this system introduces a 6×His tag to ERG10, facilitating its purification and isolation.
BBa_25KA49Y2	ADH1 Promotor-Myc-linker-Erg13-linker-PTS1	This composite part enables the localization of ERG13 within peroxisomes, thereby utilizing acetoacetyl-CoA generated by peroxisomal acetoacetyl-CoA thiolase from acetyl-CoA. ERG13 then catalyzes the conversion of acetoacetyl-CoA into HMG-CoA, which serves as a precursor of the MVA pathway. In addition, this system introduces a Myc tag to ERG13, facilitating its purification, isolation, and identification.
BBa_25EC2ES0	TEF1 Promotor-6×His-linker-HMG1-linker-PTS1	This composite part enables the localization of HMG1 within peroxisomes, thereby utilizing HMG-CoA generated by peroxisomal ERG13 from acetoacetyl-CoA. HMG1 catalyzes the conversion of HMG-CoA into MVA. MVA can diffuse out of the peroxisome into the cytosol, where it is further converted into the ginsenoside precursor 2,3-OSQ by cytosolic enzymes of S. cerevisiae, thus channeling peroxisome-derived fatty acids into ginsenoside biosynthesis. In addition, this system introduces a 6×His tag to HMG1, facilitating its purification, isolation, and identification.
BBa_25SJN43R	ADH1 Promotor-βAS-5×Leu-ADH1 terminator	This composite part enables the expression of β-amyrin synthase from Glycyrrhiza glabra in S. cerevisiae, thereby catalyzing the cyclization of 2,3-oxidosqualene into β-amyrin, which is crucial for the production of oleanolic acid (OA), a key precursor for the biosynthesis of ginsenoside Ro.
BBa_25DWGYOM	ADH1 Promotor-CYP716A12-5×Leu-ADH1 terminator	This composite part enables the expression of cytochrome P450 monooxygenase from Medicago truncatula in S. cerevisiae, thereby catalyzing the oxidation of β-amyrin to oleanolic acid, which is crucial for supplying calenduloside E (CE) as a precursor for the biosynthesis of ginsenoside Ro.
BBa_253Q8KMD	ADH1 Promotor-MtCPR-6×His-ADH1 terminator	This composite part enables the expression of Cytochrome P450 reductase 1 from Medicago truncatula isolate MTR1 in S. cerevisiae, thereby catalyzing the oxidation of β-amyrin to oleanolic acid, which is crucial for supplying calenduloside E (CE) as a precursor for the biosynthesis of ginsenoside Ro.In addition, the 6×His tag facilitates the purification and separation of MtCPR.
BBa_259T553U	ADH1 promotor-UGT73P40-ADH1 terminator	This system enables the expression of the glycosyltransferase UGT73P40 from Trifolium subterraneum in the cytosol of S. cerevisiae, where it catalyzes the transfer of a glucosyl moiety from UDP-Glc onto calenduloside E to form zingibroside R1, or onto chikusetsusaponin IVa to produce ginsenoside Ro.
BBa_25YRT715	TEF1 promotor-UGT73F3-6×His-CYC1 terminator	This system enables the expression of the glycosyltransferase UGT73F3 from Medicago truncatula in the cytosol of S. cerevisiae, where it catalyzes the transfer of a glucosyl moiety from UDP-Glc onto zingibroside R1 to form ginsenoside Ro, or onto calenduloside E to yield chikusetsusaponin IVa.In addition, this system introduces a 6×His tag to UGT73F3, facilitating its purification, isolation, and identification.
BBa_25ASXMNS	GPM1 promotor-Pn022859-6×His	This system enables the expression of the glycosyltransferase Pn022859 from Panax notoginseng in the cytosol of S. cerevisiae, where it catalyzes the transfer of a glucuronosyl moiety from UDP-GlcA onto oleanolic acid to form calenduloside E, a precursor of ginsenoside Ro. In addition, this system introduces a 6×His tag to Pn022859, facilitating its purification, isolation, and identification.
BBa_25CRW9M2	ADH1 promotor-FBPase1-6×His-CYC1 terminator	This system enables the overexpression of fructose-1,6-bisphosphatase in S. cerevisiae, where it hydrolyzes fructose-1,6-bisphosphate (FBP) to fructose-6-phosphate, providing a precursor for UDP-glucose (UDPG) biosynthesis. In addition, this system introduces a 6×His tag to fructose-1,6-bisphosphatase, facilitating its purification, isolation, and identification.
BBa_25TH9UQB	ADH1 promotor-PGM2-6×His-CYC1 terminator	This system enables the overexpression of phosphoglucomutase in S. cerevisiae, where it catalyzes the reversible conversion between glucose-1-phosphate (G-1-P) and glucose-6-phosphate (Glc-6-P), providing a precursor for UDP-glucose (UDPG) biosynthesis. In addition, this system introduces a 6×His tag to phosphoglucomutase, facilitating its purification, isolation, and identification.
BBa_25EA3WQF	TEF1 promotor-UGP1-myc	This system enables the overexpression of UTP–glucose-1-phosphate uridylyltransferase (UGP1) in S. cerevisiae, where it catalyzes the formation of UDP-glucose (UDPG) from glucose-1-phosphate (G-1-P) and UTP. In addition, this system introduces a Myc tag to UGP1, facilitating its purification, isolation, and identification.