Plasmids

From Markus Heitzer, University of Regensburg-Germany, November 2007

codon-adapted Gaussia princes luciferase under control of HSP70A/RBCS2 promoter

host strain: Pir1 (Invitrogen)
kanamycin resistant

Sequence

Fuhrmann M, Hausherr A, Ferbitz L, Schödl T, Heitzer M, Hegemann P (2004) Monitoring dynamic expression of nuclear genes in Chlamydomonas reinhardtii by using a synthetic luciferase reporter gene. Plant Mol Biol 55:869-881 Heitzer M, Zschoernig B (2007) Construction of modular tandem expression vectors for the green alga Chlamydomonas reinhardtii using the Cre/lox-system. Biotechniques 43:324

From Markus Heitzer, University of Regensburg-Germany, November 2007

loxP, HSP70A/RBCS2 promoter, codon-adapted human erythropoietin with ARS2 leader and RBCS2 introns, His6 tag, RBCS2 3′ UTR

host strain: Pir1 (Invitrogen)
kanamycin resistant

Sequence

Fuhrmann M, Hausherr A, Ferbitz L, Schödl T, Heitzer M, Hegemann P (2004) Monitoring dynamic expression of nuclear genes in Chlamydomonas reinhardtii by using a synthetic luciferase reporter gene. Plant Mol Biol 55:869-881 Heitzer M, Zschoernig B (2007) Construction of modular tandem expression vectors for the green alga Chlamydomonas reinhardtii using the Cre/lox-system. Biotechniques 43:324

From Markus Heitzer, University of Regensburg-Germany, November 2007

ARS2 promoter, codon-adapted crluc Renilla luciferase with RBCS2 introns, Hs6 tag; kanamycin resistant

host strain: Pir1 (Invitrogen)
kanamycin resistant

Sequence

Fuhrmann M, Hausherr A, Ferbitz L, Schödl T, Heitzer M, Hegemann P (2004) Monitoring dynamic expression of nuclear genes in Chlamydomonas reinhardtii by using a synthetic luciferase reporter gene. Plant Mol Biol 55:869-881

From Markus Heitzer, University of Regensburg-Germany, November 2007

ARG7.8 genomic sequence inserted into pUCBM20, followed by loxP site

host strain: DH5 alpha
amp resistant

Sequence

Heitzer M, Zschoernig B (2007) Construction of modular tandem expression vectors for the green alga Chlamydomonas reinhardtii using the Cre/lox-system. Biotechniques 43:324

From Markus Heitzer, University of Regensburg-Germany, November 2007

expression vector containing loxP, beta-tubulin promoter, aphVIII, and 3′ UTR from chlamyopsin-1

host strain: DH5 alpha
amp resistant

Sequence

Heitzer M, Zschoernig B (2007) Construction of modular tandem expression vectors for the green alga Chlamydomonas reinhardtii using the Cre/lox-system. Biotechniques 43:324

From Markus Heitzer, University of Regensburg-Germany, November 2007

Expression vector containing loxP, beta-tubulin promoter, aph7 with intron 1 from RBCS2, and 3′ UTR from chlamyopsin-1

host strain: DH5 alpha
amp resistant

Sequence

Heitzer M, Zschoernig B (2007) Construction of modular tandem expression vectors for the green alga Chlamydomonas reinhardtii using the Cre/lox-system. Biotechniques 43:324

PSAD gene, from which coding region can be removed and replaced with a gene of interest under control of the PSAD flanking sequences

host strain: DH5 alpha
amp resistant

Fischer N, Rochaix JD (2001) The flanking regions of PsaD drive efficient gene expression in the nucleus of the green alga Chlamydomonas reinhardtii. Mol Gen Genet 265:888-894

From Pete Lefebvre, University of Minnesota

plasmid used in preparing the BAC library

chloramphenicol resistant

From Xingshan Jiang, David Stern lab, Boyce Thompson Institute, August 2008

Gateway derivative of Maa7/XIR

amp resistant

Instructions

Map

From Xingshan Jiang, David Stern lab, Boyce Thompson Institute, August 2008

Gateway derivative of Maa7/XIR

amp resistant

Instructions

Map

Herbicide resistant ALS gene

This plasmid can be used as a dominant selectable marker on sulfometuron methyl (SMM) in Chlamydomonas and confers ampicillin resistance in E. coli.

host strain: DH5 alpha
amp resistant

Sequence

Kovar JL, Zhang J, Funke RP, Weeks DP (2002) Molecular analysis of the acetolactate synthase gene of Chlamydomonas reinhardtii and development of a genetically engineered gene as a dominant selectable marker for genetic transformation. Plant Journal 29:109–117

From Attila Molnar, David Baulcombe lab, University of Cambridge, December 2008

Artificial mRNAi construct

host strain: DH5 alpha
amp resistant

Sequence PDF

Sequence (.gb file)

Molnar A, Bassett A, Thuenemann E, Schwach F, Karkare S, Ossowski S, Weigel D, Baulcombe D (2009) Highly specific gene silencing by artificial microRNAs in the unicellular alga Chlamydomonas reinhardtii. Plant J 58:165-174

From Attila Molnar, David Baulcombe lab, University of Cambridge, December 2008

Artificial mRNAi construct

host strain: DH5 alpha
amp resistant

Sequence PDF

Sequence (.gb file)

Molnar A, Bassett A, Thuenemann E, Schwach F, Karkare S, Ossowski S, Weigel D, Baulcombe D (2009) Highly specific gene silencing by artificial microRNAs in the unicellular alga Chlamydomonas reinhardtii. Plant J 58:165-174

From Attila Molnar, David Baulcombe lab, University of Cambridge, December 2008

Artificial mRNAi construct

host strain: DH5 alpha
amp resistant

Sequence PDF

Sequence (.gb file)

Molnar A, Bassett A, Thuenemann E, Schwach F, Karkare S, Ossowski S, Weigel D, Baulcombe D (2009) Highly specific gene silencing by artificial microRNAs in the unicellular alga Chlamydomonas reinhardtii. Plant J 58:165-174

From Dudley Page, Merchant lab, UCLA, March 2009

This plasmid is a Gateway-adapted form of the arylsulfatase reporter vector pJD100.

host strain: Survival T1 (Invitrogen)
amp resistant

Sequence (.txt)

Sequence PDF

Schematic (.doc)

Schematic PDF

Allen MD, Kropat J, Merchant SS (2008) Regulation and localization of isoforms of the aerobic oxidative cyclase in Chlamydomonas reinhardtii. Photochem Photobiol 84:1336-42

From Wolfgang Mages, University of Regensburg, November 2009

Hygromycin resistance vector

host strain: DH5 alpha
ampicillin resistant in E. coli

Sequence

Berthold et al. 2002. Protist, Vol. 153, 401–412

From Wolfgang Mages, University of Regensburg, November 2009

Hygromycin resistance vector

This plasmid is almost identical pHyg3, only the rbcS2 intron1-sequence is absent in pHyg4. The pHyg4 sequence could easily be made by deleting the intron sequence from pHyg3.

host strain: DH5 alpha
ampicillin resistant in E. coli

pHyg3 sequence

From Michael Schroda, Max Planck Institute of Molecular Plant Physiology, August 2010

ble gene behind HSP70A-RBCS2 promoter

host strain: DH5 alpha
amp resistant

Map

Sequence (.seq file)

From Michael Schroda, Max Planck Institute of Molecular Plant Physiology, August 2010

NIT1, amiRNA, RPL12 UTR, C_310026 UTR

host strain: DH5 alpha
amp resistant

Sequence (.docx file)

Map

From Michael Schroda, Max Planck Institute of Molecular Plant Physiology, via Lai-Wa Tam, U of MN, August 2010

HSP70B gene behind HSP70A promoter

host strain: DH10B
amp resistant

Map

Sequence (.seq file)

From Michael Schroda, Max Planck Institute of Molecular Plant Physiology, via Lai-Wa Tam, U of MN, August 2010

HSP70B gene behind HSP70A-RBCS2 promoter

host strain: DH5 alpha
amp resistant

Map

Sequence (.seq file)

From Michael Schroda, Max Planck Institute of Molecular Plant Physiology, via Lai-Wa Tam, U of MN, August 2010

aadA gene behind HSP70A-RBCS2 promoter, 3’RBCS2

host strain: DH5 alpha
amp resistant

Map

Sequence (.seq file)

From Michael Schroda, Max Planck Institute of Molecular Plant Physiology, October 2010

host strain: DH5 alpha
amp resistant

Sequence (.pdw file)

From Robert Spreitzer, Univ. of Nebraska, May 2011

host strain: DH5 alpha
amp resistant

From Wenqiang Yang, Carnegie Institution, Grossman Lab, July 2011

The psaD promoter is cloned between KpnI and XhoI, and the psaD terminator between XbaI and NotI. This promoter drives the expression of the AphVIII coding sequence cloned between NdeI and EcoRV.

host strain: DH5 alpha
amp resistant

Sequence (PDF)

Sequence (.doc)

Source: Olivier Vallon, Institut de Biologie Physico-Chimique, Paris, November 2011

Used for insertional mutagenesis, complementation by cloned genomic fragments

Insert: CrAadA (AR promoter, RBCS2 3′-UTR)

Origin: CrAadA into pMS188 (AatII)

Vector: Bluescript

Host strain: DH5α

Ap resistant

Sequence (PDF)

Sequence (.txt)

Source: Olivier Vallon, Institut de Biologie Physico-Chimique, Paris, November 2011

Used for insertional mutagenesis, marker rescue

Insert: CrSpAadA (AR promoter, RBCS2 3′-UTR)

Origin: E. coli artificial promoter into pALM32 (FspAI)

Vector: Bluescript

Host strain: DH5α

Ap Sp resistant

Sequence (PDF)

Sequence (.txt)

Source: Olivier Vallon, Institut de Biologie Physico-Chimique, Paris, November 2011

Used to modify Chlamydomonas genomic BACs to Pm resistance

Insert: AphVIII (AR promoter, RBCS2 3′-UTR)

Origin: pCVD442 with IS1 and CmR removed (NdeI AsiSI) and replaced by bla, then inserting 3 kbp of pBeloBAC11-Nit8 (AscI AgeI), then inserting (FseI) the AphVIII marker from pBC1

Vector: R6K

Host strain: S17-1 λpir

Ap Pm resistant

Sequence (PDF)

Sequence (.txt)

Source: Olivier Vallon, Institut de Biologie Physico-Chimique, Paris, November 2011

Used to modify Chlamydomonas genomic BACs to Sp resistance

Insert: CrSpAadA (AR promoter, RBCS2 3′-UTR)

Origin: pCVD442 with IS1 and CmR removed (NdeI AsiSI) and replaced by bla, then inserting 3 kbp of pBeloBAC11-Nit8 (AscI AgeI), then inserting (FseI) the CrEcAadA marker from pALM33

Vector: R6K

Host strain: S17-1 λpir

Ap Sp resistant

Sequence (PDF)

Sequence (.txt)

Source: Olivier Vallon, Institut de Biologie Physico-Chimique, Paris, November 2011

Used to modify Chlamydomonas genomic BACs to Sp resistance

Insert: CrAadA (AR promoter, RBCS2 3′-UTR)

Origin: pCVD442 with IS1 and CmR removed (NdeI AsiSI) and replaced by bla, then inserting 3 kbp of pBeloBAC11-Nit8 (AscI AgeI), then inserting (FseI) the CrAadA marker from pALM32 (sense with NIT8)

Vector: R6K

Host strain: S17-1 λpir

Amp resistant

Sequence (PDF)

Sequence (.txt)