From Moritz Meyer, Martin Jonikas lab, Princeton University, August 2020

This strain was created by transforming CMJ030 with pMJ504 and is paromomycin resistant.

pMJ504: pLM005-Cre12.g498550-Venus-3xFLAG

MPM1 (Cre12.g498550) gDNA coding sequence, spanning the start codon to the triplet just before the stop codon, was cloned in frame into pLM005 by Gibson assembly (as described in https://doi.org/10.1073/pnas.1522866113.

From Moritz Meyer, Martin Jonikas lab, Princeton University, August 2020

This strain was created by transforming CMJ030 with pMJ505 and is paromomycin resistant.

pMJ505: pLM005-Cre12.g498550-Venus-3xFLAG-3xCtermSAGA2

pMJ504, containing the gDNA of Cre12.g498550, was reengineered by restriction digestion and T4 ligation to add in-frame immediately after the 3xFLAG a synthetic fragment containing three copies of the sequence coding for the 15 C-terminal amino acids of SAGA2 (Cre 09.g394621), interspersed with a short flexible linker (GGGGGS).

From Moritz Meyer, Martin Jonikas lab, Princeton University, August 2020

This strain was created by transforming CMJ030 with pMJ506 and is paromomycin resistant.

pMJ506: pLM005-Cre17.g724300-Venus-3xFLAG-3xCtermSAGA2

PSAK (Cre17.g724300) gDNA coding sequence, spanning the start codon to the triplet just before the stop codon, had been previously cloned in frame into pLM005 by Gibson assembly (as described in https://doi.org/10.1073/pnas.1522866113; pMJ280 = pLM005-Cre17.g724300-Venus-3xFLAG). That plasmid was then reengineered by restriction digestion and T4 ligation to add in-frame immediately after the 3xFLAG a synthetic fragment containing three copies of the sequence coding for the 15 C-terminal amino acids of SAGA2 (Cre 09.g394621), interspersed with a short flexible linker (GGGGGS)

From Moritz Meyer, Martin Jonikas lab, Princeton University, August 2020

This strain was created by transforming CMJ030 with pMJ507 and is paromomycin resistant.

pMJ507: pLM005-Cre10.g430350-Venus-3xFLAG

Cre10.g430350 gDNA coding sequence, spanning the start codon to the triplet just before the stop codon, was cloned in frame into pLM005 by Gibson assembly (as described in https://doi.org/10.1073/pnas.1522866113).

From Moritz Meyer, Martin Jonikas lab, Princeton University, August 2020

This strain was created by transforming CMJ030 with pMJ508 and is paromomycin resistant.

pMJ508: pLM005-Cre10.g430350[W51A/R52A]-Venus-3xFLAG

Previously synthesized pMJ507 (CC-5652) was modified by side-directed mutagenesis. Triplet coding for Tryptophan in position 51 of the mature protein now codes for an Alanine. Triplet coding for Arginine in position 52 of the mature protein now codes for an Alanine.

From Moritz Meyer, Martin Jonikas lab, Princeton University, August 2020

This strain was created by transforming CMJ030 with pMJ509 and is paromomycin resistant.

pMJ509: pLM005-Cre10.g430350[W47A/R48A/W51A/R52A/W110A/R111A/W148A/R149A]-Venus-3xFLAG

Previously synthesized pMJ507 was modified by side-directed mutagenesis. All four triplet pairs coding for a Tryptophan-Arginine dipeptide were mutagenized to code for a double Alanine (W47A/R48A/W51A/R52A/W110A/R111A/W148A/R149A).

From Moritz Meyer, Martin Jonikas lab, Princeton University, August 2020

This strain was created by transforming CMJ030 with pLM154 and is paromomycin resistant.

pLM154: pLM099-Cre09.g394621-Venus-3xFLAG

SAGA2 (Cre09.g394621) coding sequence plus 2216 bp upstream of the start codon was cloned into pLM099 by recombineering to produce a C-terminal CrVenus-3xFLAG tag.


https://www.biorxiv.org/content/10.1101/2020.08.16.252858v1.full

From Dr. Vinzenz Bayro-Kaiser, Tel Aviv University-Israel, August 2020

Mutant allele: Cre02.g076250.t1.1:c.2044_2045delCCinsTT
Background: 1A+ (137c) obtained from Prof. Jean-David Rochaix
Origin: UV induced mutagenesis followed by 5 times backcrossing to the background strain and selection
Culture maintenance: TAP media, 25 °C, 100 uEm-2s-1


Bayro-Kaiser V, Nelson N (2016)Temperature-sensitive PSII: a novel approach for sustained photosynthetic hydrogen production. Photosynth Res. 130:113-121

Bayro-Kaiser V, Nelson N (2020) Temperature Sensitive Photosynthesis: Point Mutated CEF-G, PRK, or PsbO Act as Temperature-Controlled Switches for Essential Photosynthetic Processes. Frontiers in Plant Science 11:1465

From Dr. Vinzenz Bayro-Kaiser, Tel Aviv University-Israel, August 2020

Mutant allele: Cre12.g554800.t1.2:c.479C>T
Background: 1A+ (137c) obtained from Prof. Jean-David Rochaix
Origin: UV induced mutagenesis followed by 5 times backcrossing to the background strain and selection
Culture maintenance: TAP media, 25 °C, 100 uEm-2s-1


Bayro-Kaiser V, Nelson N (2016)Temperature-sensitive PSII: a novel approach for sustained photosynthetic hydrogen production. Photosynth Res. 130:113-121

Bayro-Kaiser V, Nelson N (2020) Temperature Sensitive Photosynthesis: Point Mutated CEF-G, PRK, or PsbO Act as Temperature-Controlled Switches for Essential Photosynthetic Processes. Frontiers in Plant Science 11:1465

From Dr. Vinzenz Bayro-Kaiser, Tel Aviv University-Israel, August 2020

Mutant alleles: Cre10.g448950.t1.1:c.71_72delCTinsTC   &   Cre10.g448950.t1.1:c.101T>C   &   Cre10.g449600.t1.1:c.365G>A
Background: 1A+ (137c) obtained from Prof. Jean-David Rochaix
Origin: UV induced mutagenesis followed by 5 times backcrossing to the background strain and selection
Culture maintenance: TAP media, 25 °C, 100 uEm-2s-1


Bayro-Kaiser V, Nelson N (2016)Temperature-sensitive PSII: a novel approach for sustained photosynthetic hydrogen production. Photosynth Res. 130:113-121

Bayro-Kaiser V, Nelson N (2020) Temperature Sensitive Photosynthesis: Point Mutated CEF-G, PRK, or PsbO Act as Temperature-Controlled Switches for Essential Photosynthetic Processes. Frontiers in Plant Science 11:1465

From Dr. Vinzenz Bayro-Kaiser, Tel Aviv University-Israel, August 2020

Mutant allele: Cre09.g396213.t1.1:c.302C>A
Background: 1A+ (137c) obtained from Prof. Jean-David Rochaix
Origin: UV induced mutagenesis followed by 5 times backcrossing to the background strain and selection
Culture maintenance: TAP media, 25 °C, 100 uEm-2s-1


Bayro-Kaiser V, Nelson N (2016)Temperature-sensitive PSII: a novel approach for sustained photosynthetic hydrogen production. Photosynth Res. 130:113-121

Bayro-Kaiser V, Nelson N (2020) Temperature Sensitive Photosynthesis: Point Mutated CEF-G, PRK, or PsbO Act as Temperature-Controlled Switches for Essential Photosynthetic Processes. Frontiers in Plant Science 11:1465

From Alfredo Kono, Martin Spalding lab, Iowa State University, July 2020

This double mutant is a product of a cross of LMJ.RY0402.191570 mt- (LCI1 mutant) with pmp1 CC‐5378 mt+ (LCIB mutant) and requires high CO2 concentrations for phototrophic growth.

Mutant alleles: LCIB, Cre10.g452800.t1.1, chromosome_10:4608866..4611901 forward; LCI1, Cre03.g162800.t1.1, chromosome_3:2885264..2887871 reverse


Kono A, Spalding MH. LCI1, a Chlamydomonas reinhardtii plasma membrane protein, functions in active CO2 uptake under low CO2. Plant J. 2020 Jun;102(6):1127-1141. doi: 10.1111/tpj.14761. Epub 2020 Apr 27. PMID: 32248584.

Li X, Zhang R, Patena W, Gang SS, Blum SR, Ivanova N, Yue R, Robertson JM, Lefebvre PA, Fitz-Gibbon ST, Grossman AR, Jonikas MC. An Indexed, Mapped Mutant Library Enables Reverse Genetics Studies of Biological Processes in Chlamydomonas reinhardtii. Plant Cell. 2016 Feb;28(2):367-87. doi: 10.1105/tpc.15.00465. Epub 2016 Jan 13. PMID: 26764374; PMCID: PMC4790863.


  • Locus:
  • LCIB, LCI1
  • Chromosome:
  • 10, 3

From Alfredo Kono, Martin Spalding lab, Iowa State University, July 2020

This double mutant is the product of a cross of LMJ.RY0402.191570 mt- (LCI1 mutant) with lcia63 mt+ CC‐5066 (LCIA mutant).

Mutant alleles: LCIA, NAR 1.2, Cre06.g309000.t1.1, chromosome_6:8604782..8607670 reverse; LCI1, Cre03.g162800.t1.1, chromosome_3:2885264..2887871 reverse


Kono A, Spalding MH. LCI1, a Chlamydomonas reinhardtii plasma membrane protein, functions in active CO2 uptake under low CO2. Plant J. 2020 Jun;102(6):1127-1141. doi: 10.1111/tpj.14761. Epub 2020 Apr 27. PMID: 32248584.

Li X, Zhang R, Patena W, Gang SS, Blum SR, Ivanova N, Yue R, Robertson JM, Lefebvre PA, Fitz-Gibbon ST, Grossman AR, Jonikas MC. An Indexed, Mapped Mutant Library Enables Reverse Genetics Studies of Biological Processes in Chlamydomonas reinhardtii. Plant Cell. 2016 Feb;28(2):367-87. doi: 10.1105/tpc.15.00465. Epub 2016 Jan 13. PMID: 26764374; PMCID: PMC4790863.

Wang Y, Spalding MH. Acclimation to very low CO2: contribution of limiting CO2 inducible proteins, LCIB and LCIA, to inorganic carbon uptake in Chlamydomonas reinhardtii. Plant Physiol. 2014 Dec;166(4):2040-50. doi: 10.1104/pp.114.248294. Epub 2014 Oct 21. PMID: 25336519; PMCID: PMC4256846.


  • Locus:
  • LCIA/NAR 1.2, LCI1
  • Chromosome:
  • 6,3

From Alfredo Kono, Martin Spalding lab, Iowa State University, July 2020

This double mutant is the product of lcia90 mt+ (CC‐5067, LCIA mutant) backcrossed five times with pmp1 21gr mt- (CC‐5375, LCIB mutant) and requires high CO2 concentrations for phototrophic growth.

Mutant alleles: LCIB, Cre10.g452800.t1.1, chromosome_10:4608866..4611901 forward; LCIA, NAR 1.2, Cre06.g309000.t1.1, chromosome_6:8604782..8607670 reverse


Kono A, Spalding MH. LCI1, a Chlamydomonas reinhardtii plasma membrane protein, functions in active CO2 uptake under low CO2. Plant J. 2020 Jun;102(6):1127-1141. doi: 10.1111/tpj.14761. Epub 2020 Apr 27. PMID: 32248584.

Wang Y, Spalding MH. Acclimation to very low CO2: contribution of limiting CO2 inducible proteins, LCIB and LCIA, to inorganic carbon uptake in Chlamydomonas reinhardtii. Plant Physiol. 2014 Dec;166(4):2040-50. doi: 10.1104/pp.114.248294. Epub 2014 Oct 21. PMID: 25336519; PMCID: PMC4256846.


  • Locus:
  • LCIB, LCIA/NAR 1.2
  • Chromosome:
  • 10,6

From George B. Witman, University of Massachusetts Medical School, August 2020

This is a double mutant made by mating ift74-2 IFT74Δ130 (Brown et al., 2015) to ift81-1 IFT81(5E) (Kubo et al., 2016). The strain expresses both IFT74Δ130 and IFT81(5E) in a background otherwise null for IFT74 and IFT81. IFT74Δ130 is a version of IFT74 lacking aa 1-130 important for the protein’s interaction with the highly acidic tail (also known as E-hook) of β-tubulin. IFT81(5E) is a version of IFT81 in which five highly conserved basic residues (K73, R75, R87, K114, and R115) in the protein’s calponin-homology domain have been replaced by glutamate to reduce or eliminate the protein’s binding to tubulin. The strain is predicted to lack nearly all intraflagellar transport of tubulin. It has a strong palmelloid phenotype and forms very short flagella with normal axonemal ultrastructure.

It was created by insertional mutagenesis of the parent strains with a fragment conferring resistance to hygromycin B (aph7”) to generate ift74-2 and ift81-1, followed by transformation with a DNA fragment encoding IFT74Δ130 or IFT81(5E), respectively, and containing a paromomycin-resistance gene as a selectable marker. The transformed strains were then crossed to generate the double mutant.

Mutant alleles: ift74-2, chromosome_1:4204163-4208957; ift81-1, chromosome_17:3362408-3368081


Brown JM, Cochran DA, Craige B, Kubo T, Witman GB. Assembly of IFT trains at the ciliary base depends on IFT74. Curr Biol. 2015 Jun 15;25(12):1583-93. doi: 10.1016/j.cub.2015.04.060. Epub 2015 Jun 4. PMID: 26051893; PMCID: PMC4482480.

Kubo T, Brown JM, Bellve K, Craige B, Craft JM, Fogarty K, Lechtreck KF, Witman GB. Together, the IFT81 and IFT74 N-termini form the main module for intraflagellar transport of tubulin. J Cell Sci. 2016 May 15;129(10):2106-19. doi: 10.1242/jcs.187120. Epub 2016 Apr 11. PMID: 27068536; PMCID: PMC5506485.

Van De Weghe JC, Harris JA, Kubo T, Witman GB, Lechtreck KF. Diffusion rather than IFT likely provides most of the tubulin required for axonemal assembly. J Cell Sci. 2020 Aug 14:jcs.249805. doi: 10.1242/jcs.249805. Epub ahead of print. PMID: 32801124.


  • Locus:
  • IFT74, IFT81
  • Chromosome:
  • 1,17

From George B. Witman, University of Massachusetts Medical School, August 2020

In this strain, IFT81-HA is incorporated into the IFT-B complex and localizes normally in cell bodies and flagella. The cells appear to be fully rescued for motility and flagellar length.

It was created by insertional mutagenesis of the parent strain with a fragment conferring resistance to hygromycin B (aph7”) to generate the null mutant ift81-1, followed by rescue with a DNA fragment encoding IFT81 with a 3xhemagglutinin (HA) tag at its C-terminal end and containing a paromomycin-resistance gene as a selectable marker.

Mutant allele: ift81-1, chromosome_17:3362408-3368081


Kubo T, Brown JM, Bellve K, Craige B, Craft JM, Fogarty K, Lechtreck KF, Witman GB. Together, the IFT81 and IFT74 N-termini form the main module for intraflagellar transport of tubulin. J Cell Sci. 2016 May 15;129(10):2106-19. doi: 10.1242/jcs.187120. Epub 2016 Apr 11. PMID: 27068536; PMCID: PMC5506485.


  • Locus:
  • IFT81
  • Chromosome:
  • 17

From George B. Witman, University of Massachusetts Medical School, August 2020

This strain expresses IFT81(2E) in a background null for IFT81. IFT81(2E) is a version of IFT81 in which two highly conserved basic residues (K73 and R75) in the protein’s calponin-homology domain, implicated in binding to tubulin, have been replaced by glutamate. The strain has apparently normal length flagella, normal flagellar regeneration kinetics, normal motility, and normal IFT. The frequency of anterograde tubulin IFT was not significantly reduced in the flagella of this strain.

It was created by insertional mutagenesis of the parent strain with a fragment conferring resistance to hygromycin B (aph7”) to generate ift81-1, followed by transformation with a DNA fragment encoding IFT81(2E) and containing a paromomycin-resistance gene as a selectable marker.

Mutant allele: ift81-1, chromosome_17:3362408-3368081


Kubo T, Brown JM, Bellve K, Craige B, Craft JM, Fogarty K, Lechtreck KF, Witman GB. Together, the IFT81 and IFT74 N-termini form the main module for intraflagellar transport of tubulin. J Cell Sci. 2016 May 15;129(10):2106-19. doi: 10.1242/jcs.187120. Epub 2016 Apr 11. PMID: 27068536; PMCID: PMC5506485.


  • Locus:
  • IFT81
  • Chromosome:
  • 17