PART COLLECTION

Our design consists of three main cycles：

Cycle 1: Construction and verification of carbohydrate utilization module.

pCOLAtrc(lost O)-sacC-aga, BBa_25TFWJDX

Cycle 2: Construction and verification of galactose self-induction module.

Part 1: Promoter Characterization

pCOM4-PgalP-a-EGFP, BBa_2572ULRB

pCOM4-PgalP-b-EGFP, BBa_25NRQBKP

pCOM4-PgalP-c-EGFP, BBa_25Q32CXO

Part 2：Substituting GFP with GalP

pCOM4-PgalP-GalP, BBa_25M3YRM6

Cycle 3: Construction and verification of 3'-SL synthesis module.

pET-CSS-SaiT, BBa_251TW3T1

In summary, the synergistic action of cycles 1, 2, and 3 is essential to enable the use of yellow serofluid as a carbon source and harness its inherent galactose utilization pathway for the production of 3'-sialyllactose (3'-SL)

Number	Name	Part type	Part type	Part function	Part diagram
BBa_25V1IYRD	SacC	Basic Part	Coding	SacC is an extracellular enzyme that primarily catalyzes the hydrolysis of large fructan polymers (such as Levan and Inulin) into fructose molecules
BBa_25MWK240	Aga	Basic Part	Coding	Alpha-galactosidase is a glycoside hydrolase that specifically catalyzes the hydrolysis of α-galactosidic bonds. It cleaves the terminal α-1,6-linked galactose residues from oligosaccharides and polysaccharides, yielding galactose and the corresponding substrate.
BBa_25K4V860	PgalP-a	Basic Part	Promoter	Mutants of PgalP were engineered to improve the initial expression level of GalP, thereby enhancing the intracellular availability of galactose
BBa_25ZG3I79	PgalP-b	Basic Part	Promoter	Mutants of PgalP were engineered to improve the initial expression level of GalP, thereby enhancing the intracellular availability of galactose
BBa_255LQDVV	PgalP-c	Basic Part	Promoter	Mutants of PgalP were engineered to improve the initial expression level of GalP, thereby enhancing the intracellular availability of galactose
BBa_25M20CQN	EGFP	Basic Part	Coding	EGFP can emit bright green fluorescence under ultraviolet light or blue light, which is convenient for researchers to track and observe various biological processes in cells, tissues or organs.
BBa_254XGI7O	galP	Basic Part	Coding	Mutants of PgalP were engineered to improve the initial expression level of GalP, thereby enhancing the intracellular availability of galactose
BBa_250SNH60	CSS	Basic Part	Coding	CSS catalyzes the formation of CMP-Neu5Ac. Under the action of CSS, free N-acetylneuraminic acid (Neu5Ac) combines with cytidine triphosphate (CTP) to form CMP-Neu5Ac, simultaneously releasing pyrophosphate (PPi)
BBa_25JU2UK2	α2,3-SiaT	Basic Part	Coding	α2,3-SiaT is responsible for transferring the Neu5Ac moiety from CMP-Neu5Ac to the galactose residue of lactose, forming an α2,3-glycosidic bond to produce 3'-SL
BBa_258YB0NG	pCOLADuet-1	Basic Part	Plasmid_Backbone	As a plasmid construction vector
BBa_25TFWJDX	pCOLAtrc(lostO)-sacC-aga	Composite Part	Plasmid	After the plasmid is transformed into E.coli, it functions, so that E.coli can survive in soy whey
BBa_2572ULRB	pCOM4-PgalP-a-EGFP	Composite Part	Plasmid	After the plasmid was transformed into Escherichia coli, it played a role. The promoter with the strongest induction ability was screened by measuring the fluorescence intensity.
BBa_25NRQBKP	pCOM4-PgalP-b-EGFP	Composite Part	Plasmid	After the plasmid was transformed into Escherichia coli, it played a role. The promoter with the strongest induction ability was screened by measuring the fluorescence intensity.
BBa_25Q32CXO	pCOM4-PgalP-c-EGFP	Composite Part	Plasmid	After the plasmid was transformed into Escherichia coli, it played a role. The promoter with the strongest induction ability was screened by measuring the fluorescence intensity.
BBa_25M3YRM6	pCOM4-PgalP-GalP	Composite Part	Plasmid	Responsible for the transport of galactose in cells and activate the expression of downstream proteins.
BBa_259IMP7D	pETDuet-1	Basic Part	Plasmid_Backbone	As a plasmid construction vector
BBa_251TW3T1	pET-CSS-SaiT	Composite Part	Plasmid	Responsible for the synthesis of 3'-SL in cells.