Coding

Part:BBa_K5218007

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


PpC3H

p-coumaric acid 3-hdroxylase (C3H) gene from Physcomitrella patensa.

Base Pairs：1601 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 EcoRI site found at 690
    Illegal XbaI site found at 369
    Illegal XbaI site found at 573
    Illegal PstI site found at 855
    Illegal PstI site found at 1539
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal EcoRI site found at 690
    Illegal PstI site found at 855
    Illegal PstI site found at 1539
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal EcoRI site found at 690
  • 23
    INCOMPATIBLE WITH RFC[23]
    Illegal EcoRI site found at 690
    Illegal XbaI site found at 369
    Illegal XbaI site found at 573
    Illegal PstI site found at 855
    Illegal PstI site found at 1539
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal EcoRI site found at 690
    Illegal XbaI site found at 369
    Illegal XbaI site found at 573
    Illegal PstI site found at 855
    Illegal PstI site found at 1539
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI.rc site found at 352
    Illegal BsaI.rc site found at 933
    Illegal BsaI.rc site found at 1197

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 PpC3H gene loci is LOC112275074, transcript loci is XM_024505055.1 (NCBI). Team BGI-MammothEdu-South 2024 extracted Physcomitrella patensa sporophyte total RNA, reverse transcribed into cDNA and cloned the PpC3H 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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