Coding

Part:BBa_K5218009

Designed by: Amy Fu, Juliette, Lichuan Chen, Xiaojuan Wang   Group: iGEM24_BGI-MammothEdu-South   (2024-09-03)

CrC3H

p-coumaric acid 3-hdroxylase (C3H) gene from Catharanthus roseus.

Base Pairs：1569 bp

Function：p-coumaric acid 3-hdroxylase that catalyzes the formation of caffeic acid from p-coumaric acid.

Figure.1: C3H plays a role in the Caffeic acid cycle.
figure adopted from Tatiana et al., 2020[1]

Sequence and Features


Assembly Compatibility:
  • 10
    INCOMPATIBLE WITH RFC[10]
    Illegal PstI site found at 1535
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal PstI site found at 1535
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    INCOMPATIBLE WITH RFC[23]
    Illegal PstI site found at 1535
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal PstI site found at 1535
  • 1000
    COMPATIBLE WITH RFC[1000]

Introduction

Cytochrome P450 family has been identified in all kingdoms of life, which can catalyze more than 20 types of reactions, including hydroxylation, epoxidation, cyclization, and so on [1]. One of its members, C3H, catalyzes the formation of caffeic acid from p-coumaric acid. This reaction is associated with lignin biosynthesis, which is vital for plant development [2]. Recently scientists have utilized Fungal Bioluminescence Pathway (FBP) to generate autoluminescent plant [3]. C3H, which produced the starting substance for caffeic acid cycle, is a crucial candidate gene for FBP genetical modification.

The engineering objective of this project is to generate brighter autoluminescent plants using synthetic biology approaches. Team BGI-MammothEdu-South 2024 selected a variety of candidate genes from different species and tested their functions in FBP via eGFP reporter system (pS1300-GFP plasmid). The CrC3H gene loci is JAMLDZ010000192.1, transcript loci is M9H77_00625 (NCBI). Team BGI-MammothEdu-South 2024 extracted Catharanthus roseus flower and young leaf total RNA, reverse transcribed into cDNA and cloned the CrC3H CDS with specific primer pairs.

Reference

1.Jung S T, Lauchli R, Arnold F H. (2011) Cytochrome P450: taming a wild type enzyme[J]. Current opinion in biotechnology, 22(6): 809-817.

2.Barros J, Escamilla-Trevino L, Song L, Rao X, Serrani-Yarce JC, Palacios MD, Engle N, Choudhury FK, Tschaplinski TJ, Venables BJ, Mittler R, Dixon RA. 4-Coumarate 3-hydroxylase in the lignin biosynthesis pathway is a cytosolic ascorbate peroxidase. (1994) Nature Communication, 2019 Apr 30;10(1).

3.Mitiouchkina, T., Mishin, A.S., Somermeyer, L.G. et al. Plants with genetically encoded autoluminescence. (2020) Nature Biotechnology, 38, 944–946.

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