In vivo assembly of DNA-fragments in the moss, Physcomitrella patens

Brian Christopher King; Konstantinos Vavitsas; Nur Kusaira Binti Khairul Ikram; Josephine Schrøder; Lars Scharff; Björn Robert Hamberger; Poul Erik Jensen; Henrik Toft Simonsen

doi:10.1038/srep25030

In vivo assembly of DNA-fragments in the moss, Physcomitrella patens

Brian Christopher King, Konstantinos Vavitsas, Nur Kusaira Binti Khairul Ikram, Josephine Schrøder, Lars Scharff, Björn Robert Hamberger, Poul Erik Jensen, Henrik Toft Simonsen

16 Citationer (Scopus)

98 Downloads (Pure)

Abstract

Direct assembly of multiple linear DNA fragments via homologous recombination, a phenomenon known as in vivo assembly or transformation associated recombination, is used in biotechnology to assemble DNA constructs ranging in size from a few kilobases to full synthetic microbial genomes. It has also enabled the complete replacement of eukaryotic chromosomes with heterologous DNA. The moss Physcomitrella patens, a non-vascular and spore producing land plant (Bryophyte), has a well-established capacity for homologous recombination. Here, we demonstrate the in vivo assembly of multiple DNA fragments in P. patens with three examples of effective genome editing: we (i) efficiently deleted a genomic locus for diterpenoid metabolism yielding a biosynthetic knockout, (ii) introduced a salt inducible promoter, and (iii) re-routed endogenous metabolism into the formation of amorphadiene, a precursor of high-value therapeutics. These proof-of-principle experiments pave the way for more complex and increasingly flexible approaches for large-scale metabolic engineering in plant biotechnology.

Originalsprog	Engelsk
Artikelnummer	25030
Tidsskrift	Scientific Reports
Vol/bind	6
Antal sider	7
ISSN	2045-2322
DOI	https://doi.org/10.1038/srep25030
Status	Udgivet - 29 apr. 2016

Adgang til dokumentet

10.1038/srep25030

In vivo assembly of DNA-fragments in the moss, Physcomitrella patensForlagets udgivne version, 758 KB

Citationsformater

@article{d0d72cec14584fc3ae68c95a74114be3,

title = "In vivo assembly of DNA-fragments in the moss, Physcomitrella patens",

abstract = "Direct assembly of multiple linear DNA fragments via homologous recombination, a phenomenon known as in vivo assembly or transformation associated recombination, is used in biotechnology to assemble DNA constructs ranging in size from a few kilobases to full synthetic microbial genomes. It has also enabled the complete replacement of eukaryotic chromosomes with heterologous DNA. The moss Physcomitrella patens, a non-vascular and spore producing land plant (Bryophyte), has a well-established capacity for homologous recombination. Here, we demonstrate the in vivo assembly of multiple DNA fragments in P. patens with three examples of effective genome editing: we (i) efficiently deleted a genomic locus for diterpenoid metabolism yielding a biosynthetic knockout, (ii) introduced a salt inducible promoter, and (iii) re-routed endogenous metabolism into the formation of amorphadiene, a precursor of high-value therapeutics. These proof-of-principle experiments pave the way for more complex and increasingly flexible approaches for large-scale metabolic engineering in plant biotechnology.",

author = "King, {Brian Christopher} and Konstantinos Vavitsas and {Binti Khairul Ikram}, {Nur Kusaira} and Josephine Schr{\o}der and Lars Scharff and Hamberger, {Bj{\"o}rn Robert} and Jensen, {Poul Erik} and Simonsen, {Henrik Toft}",

year = "2016",

month = apr,

day = "29",

doi = "10.1038/srep25030",

language = "English",

volume = "6",

journal = "Scientific Reports",

issn = "2045-2322",

publisher = "nature publishing group",

}

TY - JOUR

T1 - In vivo assembly of DNA-fragments in the moss, Physcomitrella patens

AU - King, Brian Christopher

AU - Vavitsas, Konstantinos

AU - Binti Khairul Ikram, Nur Kusaira

AU - Schrøder, Josephine

AU - Scharff, Lars

AU - Hamberger, Björn Robert

AU - Jensen, Poul Erik

AU - Simonsen, Henrik Toft

PY - 2016/4/29

Y1 - 2016/4/29

N2 - Direct assembly of multiple linear DNA fragments via homologous recombination, a phenomenon known as in vivo assembly or transformation associated recombination, is used in biotechnology to assemble DNA constructs ranging in size from a few kilobases to full synthetic microbial genomes. It has also enabled the complete replacement of eukaryotic chromosomes with heterologous DNA. The moss Physcomitrella patens, a non-vascular and spore producing land plant (Bryophyte), has a well-established capacity for homologous recombination. Here, we demonstrate the in vivo assembly of multiple DNA fragments in P. patens with three examples of effective genome editing: we (i) efficiently deleted a genomic locus for diterpenoid metabolism yielding a biosynthetic knockout, (ii) introduced a salt inducible promoter, and (iii) re-routed endogenous metabolism into the formation of amorphadiene, a precursor of high-value therapeutics. These proof-of-principle experiments pave the way for more complex and increasingly flexible approaches for large-scale metabolic engineering in plant biotechnology.

AB - Direct assembly of multiple linear DNA fragments via homologous recombination, a phenomenon known as in vivo assembly or transformation associated recombination, is used in biotechnology to assemble DNA constructs ranging in size from a few kilobases to full synthetic microbial genomes. It has also enabled the complete replacement of eukaryotic chromosomes with heterologous DNA. The moss Physcomitrella patens, a non-vascular and spore producing land plant (Bryophyte), has a well-established capacity for homologous recombination. Here, we demonstrate the in vivo assembly of multiple DNA fragments in P. patens with three examples of effective genome editing: we (i) efficiently deleted a genomic locus for diterpenoid metabolism yielding a biosynthetic knockout, (ii) introduced a salt inducible promoter, and (iii) re-routed endogenous metabolism into the formation of amorphadiene, a precursor of high-value therapeutics. These proof-of-principle experiments pave the way for more complex and increasingly flexible approaches for large-scale metabolic engineering in plant biotechnology.

U2 - 10.1038/srep25030

DO - 10.1038/srep25030

M3 - Journal article

C2 - 27126800

SN - 2045-2322

VL - 6

JO - Scientific Reports

JF - Scientific Reports

M1 - 25030

ER -

In vivo assembly of DNA-fragments in the moss, Physcomitrella patens

Abstract

Adgang til dokumentet

Fingeraftryk

Citationsformater