Strains

From Duanmu Deqiang, Lagarias lab, University of California Davis, 8/22/13

The mutant was first created in the Niyogi lab at UC Berkeley. At UC Davis, the original mutant was first crossed to CC-621 (137c mt-) and mt- progeny was then backcrossed four times to 4A+ (CC-4051). The mutant is resistant to paromomycin (20 μg/mL).

This hmox1 mutant was then complemented by expression of the wild type HMOX1 genomic DNA under control of constitutive promoter (HSP70RbcS2). The complementing line is resistant to both paromomycin (hmox1 mutant allele) and Zeocin (the overexpression construct).

Duanmu D, Casero D, Dent RM, Gallaher S, Yang W, Rockwell NC, Martin SS, Pellegrini M, Niyogi KK, Merchant SS, Grossman AR, Lagarias JC (2013) Retrograde bilin signaling enables Chlamydomonas greening and phototrophic survival. Proc Natl Acad Sci U S A 110:3621-3626

  • Locus:
  • HMOX
  • Chromosome:
  • 10

From Duanmu Deqiang, Lagarias lab, University of California Davis, 8/22/13

This double mutant was derived by crossing the single mutants hmox1 and hmox2 and it is resistant to paromomycin (20 μg/mL).

Compared to the hmox1 or hmox2 single mutant, the double mutant has reduced capability of using extracellular heme as iron source under iron limiting conditions.

Duanmu D, Casero D, Dent RM, Gallaher S, Yang W, Rockwell NC, Martin SS, Pellegrini M, Niyogi KK, Merchant SS, Grossman AR, Lagarias JC (2013) Retrograde bilin signaling enables Chlamydomonas greening and phototrophic survival. Proc Natl Acad Sci U S A 110:3621-3626

  • Locus:
  • HMOX1 HMOX2
  • Chromosome:
  • 10,11

From Duanmu Deqiang, Lagarias lab, University of California Davis, 8/22/13

This double mutant was derived by crossing the single mutants hmox1 and hmox2 and it is resistant to paromomycin (20 μg/mL).

Compared to the hmox1 or hmox2 single mutant, the double mutant has reduced capability of using extracellular heme as iron source under iron limiting conditions.

Duanmu D, Casero D, Dent RM, Gallaher S, Yang W, Rockwell NC, Martin SS, Pellegrini M, Niyogi KK, Merchant SS, Grossman AR, Lagarias JC (2013) Retrograde bilin signaling enables Chlamydomonas greening and phototrophic survival. Proc Natl Acad Sci U S A 110:3621-3626

  • Locus:
  • HMOX1 HMOX2
  • Chromosome:
  • 10,11

From Duanmu Deqiang, Lagarias lab, University of California Davis, 8/22/13

The mutant was first created in the Niyogi lab at UC Berkeley. At UC Davis, the original mutant was first crossed to CC-621 (137c mt-) and mt- progeny was then backcrossed four times to 4A+ (CC-4051). The mutant is resistant to paromomycin (20 μg/mL).

The HMOX1 is targeted to the chloroplast and catalyzes the degradation of heme to biliverdin, which is then further reduced to phycocyanobilin by bilin reductase (PCYA, also localized to the chloroplast). Phycocyanobilin could be a retrograde signal that coordinates nuclear gene expression changes for light adaptation during dark to light transition in Chlamydomonas. Such chloroplast-nucleus signaling role of bilin is possibly also present in other photosynthetic organisms including higher plants.

This mutant is a null mutant. The plasmid insert is in the first exon of the gene. HMOX1 protein was not expressed in the mutant based on Western analysis.

Duanmu D, Casero D, Dent RM, Gallaher S, Yang W, Rockwell NC, Martin SS, Pellegrini M, Niyogi KK, Merchant SS, Grossman AR, Lagarias JC (2013) Retrograde bilin signaling enables Chlamydomonas greening and phototrophic survival. Proc Natl Acad Sci U S A 110:3621-3626

  • Locus:
  • HMOX1
  • Chromosome:
  • 10

From Duanmu Deqiang, Lagarias lab, University of California Davis, 8/22/13

This mutant was first created in the Grossman lab at Stanford using CC-4425 cw15 nit2-203 mt+ [D66]. At UC Davis, the original mutant was first crossed to CC-621 (137c mt-) and mt- progeny was then backcrossed four times to 4A+ (CC-4051). The mutant is resistant to paromomycin (20 μg/mL).

The HMOX2 is localized to the cytosol (possibly associated to the ER membrane) and catalyzes the degradation of heme to biliverdin. The HMOX2 is required for iron acquisition from heme under iron limiting conditions. This animal type heme oxygenase is present in many Chlorophyte algae but absent in Streptophyte or plants. The plasmid insert is in the last exon of the gene. The transcript abundance of the mutant was reduced to around 20% of wild type.

Duanmu D, Casero D, Dent RM, Gallaher S, Yang W, Rockwell NC, Martin SS, Pellegrini M, Niyogi KK, Merchant SS, Grossman AR, Lagarias JC (2013) Retrograde bilin signaling enables Chlamydomonas greening and phototrophic survival. Proc Natl Acad Sci U S A 110:3621-3626

  • Locus:
  • HMOX2
  • Chromosome:
  • 11

From Krishna Niyogi, University of California Berkeley, 9/12/13

This is a near-isogenic strain generated by 5 backcrosses of 17D− (CC-2191) to 4A+ (CC-4051), which showed similar sensitivity to high light and reactive oxygen species generators as 4A+ (CC-4051).

Dent RM, Haglund CM, Chin BL, Kobayashi MC, Niyogi KK (2005) Functional genomics of eukaryotic photosynthesis using insertional mutagenesis of Chlamydomonas reinhardtii. Plant Physiol 137:545-556

From Krishna Niyogi, University of California Berkeley, 9/12/13

This strain is more tolerant to singlet oxygen-producing chemicals and shows a constitutively higher expression of glutathione peroxidase (GPXH) and glutathione S-transferase (GSTS1).

Fischer BB, Ledford HK, Wakao S, Huang SG, Casero D, Pellegrini M, Merchant SS, Koller A, Eggen RI, Niyogi KK (2012) SINGLET OXYGEN RESISTANT 1 links reactive electrophile signaling to singlet oxygen acclimation in Chlamydomonas reinhardtii. Proc Natl Acad Sci U S A 109:E1302-11

  • Locus:
  • SOR1
  • Chromosome:
  • 7

From Krishna Niyogi, University of California Berkeley, 9/12/13

This strain is more tolerant to singlet oxygen-producing chemicals and shows a constitutively higher expression of glutathione peroxidase (GPXH) and glutathione S-transferase (GSTS1).

Fischer BB, Ledford HK, Wakao S, Huang SG, Casero D, Pellegrini M, Merchant SS, Koller A, Eggen RI, Niyogi KK (2012) SINGLET OXYGEN RESISTANT 1 links reactive electrophile signaling to singlet oxygen acclimation in Chlamydomonas reinhardtii. Proc Natl Acad Sci U S A 109:E1302-11

  • Locus:
  • SOR1
  • Chromosome:
  • 7

From Krishna Niyogi, University of California Berkeley, 9/12/13

Constitutive overexpression of glutathione peroxidase (GPXH) under control of PSAD promoter in pSL18.

Ledford HK, Chin BL, Niyogi KK (2007) Acclimation to singlet oxygen stress in Chlamydomonas reinhardtii. Eukaryot Cell 6:919-30

From Krishna Niyogi, University of California Berkeley, 9/12/13

Constitutive overexpression of glutathione peroxidase (GPXH) under control of PSAD promoter in pSL18.

Ledford HK, Chin BL, Niyogi KK (2007) Acclimation to singlet oxygen stress in Chlamydomonas reinhardtii. Eukaryot Cell 6:919-30

From Krishna Niyogi, University of California Berkeley, 9/12/13

Constitutive overexpression of glutathione peroxidase (GPXH) under control of PSAD promoter in pSL18.

Ledford HK, Chin BL, Niyogi KK (2007) Acclimation to singlet oxygen stress in Chlamydomonas reinhardtii. Eukaryot Cell 6:919-30

From Krishna Niyogi, University of California Berkeley, 9/12/13

Constitutive overexpression of glutathione S-transferase (GSTS1) under control of PSAD promoter in pSL18.

Ledford HK, Chin BL, Niyogi KK (2007) Acclimation to singlet oxygen stress in Chlamydomonas reinhardtii. Eukaryot Cell 6:919-30

From Krishna Niyogi, University of California Berkeley, 9/12/13

Constitutive overexpression of glutathione S-transferase (GSTS1) under control of PSAD promoter in pSL18.

Ledford HK, Chin BL, Niyogi KK (2007) Acclimation to singlet oxygen stress in Chlamydomonas reinhardtii. Eukaryot Cell 6:919-30

From Krishna Niyogi, University of California Berkeley, 9/12/13

Constitutive overexpression of glutathione S-transferase (GSTS1) under control of PSAD promoter in pSL18.

Ledford HK, Chin BL, Niyogi KK (2007) Acclimation to singlet oxygen stress in Chlamydomonas reinhardtii. Eukaryot Cell 6:919-30

From Krishna Niyogi, University of California Berkeley, 9/12/13

This vte3 mutant contains a mutation in the homolog of a 2-methyl-6-phytyl-1,4-benzoquinone methyltransferase gene found in plants.

Sirikhachornkit A, Shin JW, Baroli I, Niyogi KK (2009) Replacement of alpha-tocopherol by beta-tocopherol enhances resistance to photooxidative stress in a xanthophyll-deficient strain of Chlamydomonas reinhardtii. Eukaryot Cell 8:1648-1657

  • Locus:
  • VTE3
  • Chromosome:
  • 14

From Krishna Niyogi, University of California Berkeley, 9/12/13

Sirikhachornkit A, Shin JW, Baroli I, Niyogi KK (2009) Replacement of alpha-tocopherol by beta-tocopherol enhances resistance to photooxidative stress in a xanthophyll-deficient strain of Chlamydomonas reinhardtii. Eukaryot Cell 8:1648-1657

  • Locus:
  • VTE3, NPQ1, LOR1
  • Chromosome:
  • 14,2

From Krishna Niyogi, University of California Berkeley, September 2013

This strain is defective in nonphotochemical quenching due to lack of LHCSR3.1 and LHCSR3.2 genes.

Niyogi KK, Bjorkman O, Grossman AR (1997) Chlamydomonas Xanthophyll Cycle Mutants Identified by Video Imaging of Chlorophyll Fluorescence Quenching. Plant Cell 9:1369-1380

Peers G, Truong TB, Ostendorf E, Busch A, Elrad D, Grossman AR, Hippler M, Niyogi KK (2009) An ancient light-harvesting protein is critical for the regulation of algal photosynthesis. Nature 462:518-21

Ballottari M, Truong TB, De Re E, Erickson E, Stella GR, Fleming GR, Bassi R, Niyogi KK (2016) Identification of pH-sensing Sites in the Light Harvesting Complex Stress-related 3 Protein Essential for Triggering Non-photochemical Quenching in Chlamydomonas reinhardtii. J Biol Chem. 291:7334-46

  • Locus:
  • NPQ4 [LHCSR3]
  • Chromosome:

From Krishna Niyogi, University of California Berkeley,September 2013

This strain is defective in nonphotochemical quenching due to lack of LHCSR3.1 and LHCSR3.2 genes.

Niyogi KK, Bjorkman O, Grossman AR (1997) Chlamydomonas Xanthophyll Cycle Mutants Identified by Video Imaging of Chlorophyll Fluorescence Quenching. Plant Cell 9:1369-1380

Peers G, Truong TB, Ostendorf E, Busch A, Elrad D, Grossman AR, Hippler M, Niyogi KK (2009) An ancient light-harvesting protein is critical for the regulation of algal photosynthesis. Nature 462:518-21

Ballottari M, Truong TB, De Re E, Erickson E, Stella GR, Fleming GR, Bassi R, Niyogi KK (2016) Identification of pH-sensing Sites in the Light Harvesting Complex Stress-related 3 Protein Essential for Triggering Non-photochemical Quenching in Chlamydomonas reinhardtii. J Biol Chem. 291:7334-46

  • Locus:
  • NPQ4 [LHCSR3]
  • Chromosome:

From George Witman, University of Massachusetts Medical School, 1/10/14

This mutant was generated by transforming ptx2-1 with plasmid pGP535 (containing the PTX2 gene) to rescue ptx2 but not the oda13 (i.e., DLC6) deletion.

The ptx2-1 strain was produced by transforming strain g1 (nit1, NIT2, agg1, mt+; derivative of crosses between CC-124 and nit1-305) with pMN24.

Pazour GJ, Witman GB (2000) Forward and reverse genetic analysis of microtubule motors in Chlamydomonas. Methods 22:285-298

DiBella LM, Gorbatyuk O, Sakato M, Wakabayashi K, Patel-King RS, Pazour GJ, Witman GB, King SM (2005) Differential light chain assembly influences outer arm dynein motor function. Mol Biol Cell 16:5661-5674

The reference for the original ptx2-1 strain is:

Pazour GJ, Sineshchekov OA, Witman GB (1995) Mutational analysis of the phototransduction pathway of Chlamydomonas reinhardtii. J Cell Biol 131:427-440

  • Locus:
  • ODA13
  • Chromosome:
  • 3

From Susan Dutcher, Washington University in St. Loius, January 2014

Lux FG 3rd, Dutcher SK (1991) Genetic interactions at the FLA10 locus: suppressors and synthetic phenotypes that affect the cell cycle and flagellar function in Chlamydomonas reinhardtii. Genetics 128:549-61

Miller MS, Esparza JM, Lippa AM, Lux FG 3rd, Cole DG, Dutcher SK (2005) Mutant kinesin-2 motor subunits increase chromosome loss. Mol Biol Cell 16:3810-20

  • Locus:
  • FLA10
  • Chromosome:
  • 7

From Susan Dutcher, Washington University in St. Loius, January 2014

Progeny from CC-124 x CC-125 was crossed with S1C5 to introduce NIT1 and NIT2 and then crossed back to CC-125 again. This strain has about ¼ S1C5 markers.

  • Locus:
  • AGG1
  • Chromosome:
  • 13

From Wenqiang Yang, Carnegie Institution for Science, Grossman Lab, February 2014

Riekhof WR, Ruckle ME, Lydic TA, Sears BB, Benning C (2003) The sulfolipids 2'-O-acyl-sulfoquinovosyldiacylglycerol and sulfoquinovosyldiacylglycerol are absent from a Chlamydomonas reinhardtii mutant deleted in SQD1. Plant Physiol 133:864-74

  • Locus:
  • NIT1
  • Chromosome:
  • 9

From Robert J. Spreitzer, University of Nebraska, March 2014

Phenotype: requires acetate, sensitive to light

This nuclear mutant was isolated by Hong and Spreitzer (1994) by screening a collection of acetate-requiring mutants for lack of complementation with a known Rubisco rbcL mutant. The mutant is specifically defective in chloroplast rbcL mRNA accumulation. This strain has been maintained with acetate medium in darkness since its isolation.

Hong S, Spreitzer RJ (1994) Nuclear mutation inhibits expression of the chloroplast gene that encodes the large subunit of ribulose-1,5-bisphosphate carboxylase/oxygenase. Plant Physiol 106:673-678

From Robert J. Spreitzer, University of Nebraska, March 2014

Phenotype: requires acetate, sensitive to light

The 76-5EN nuclear mutant was isolated by Hong and Spreitzer (1994) by screening a collection of acetate-requiring mutants for lack of complementation with a known Rubisco rbcL mutant. The mutant was crossed with pf2 mt- to confirm mendelian inheritance, and this mt+ progeny clone was retained for analysis. The mutant is specifically defective in chloroplast rbcL mRNA accumulation. This strain has been maintained with acetate medium in darkness since its isolation.

Hong S, Spreitzer RJ (1994) Nuclear mutation inhibits expression of the chloroplast gene that encodes the large subunit of ribulose-1,5-bisphosphate carboxylase/oxygenase. Plant Physiol 106:673-678

From Robert J. Spreitzer, University of Nebraska, March 2014

Phenotype: requires acetate, sensitive to light

The 76-5EN nuclear mutant was isolated by Hong and Spreitzer (1994) by screening a collection of acetate-requiring mutants for lack of complementation with a known Rubisco rbcL mutant. The mutant was crossed with pf2 mt- to confirm mendelian inheritance, and this mt- progeny clone was retained for further genetic analysis. The mutant is specifically defective in chloroplast rbcL mRNA accumulation. This strain has been maintained with acetate medium in darkness since its isolation.

Hong S, Spreitzer RJ (1994) Nuclear mutation inhibits expression of the chloroplast gene that encodes the large subunit of ribulose-1,5-bisphosphate carboxylase/oxygenase. Plant Physiol 106:673-678

From Robert J. Spreitzer, University of Nebraska, March 2014

Phenotype: requires acetate, sensitive to light

The 76-5EN nuclear mutant was isolated by Hong and Spreitzer (1994) by screening a collection of acetate-requiring mutants for lack of complementation with a known Rubisco rbcL mutant. The mutant was crossed with pf2 mt- to confirm mendelian inheritance, and this pf2 mt- progeny clone was retained for further genetic analysis. The mutant is specifically defective in chloroplast rbcL mRNA accumulation. This strain has been maintained with acetate medium in darkness since its isolation.

Hong S, Spreitzer RJ (1994) Nuclear mutation inhibits expression of the chloroplast gene that encodes the large subunit of ribulose-1,5-bisphosphate carboxylase/oxygenase. Plant Physiol 106:673-678

  • Locus:
  • PF2
  • Chromosome:
  • 11

From Robert J. Spreitzer, University of Nebraska, March 2014

This strain was recovered from a cross between R107-4A mt+ and 76-5EN pf2 mt- (Hong 1996). It has been maintained with acetate medium in darkness since its isolation. The R107-4A photosynthesis-competent revertant was selected from the Rubisco rbcL-mRNA deficient 76-5EN nuclear mutant (Hong and Spreitzer 1994, 1998). When R107-4A mt+ was crossed with “wild-type” mt-, only wild-type progeny were obtained (Hong and Spreitzer 1998). When R107-4A mt+ was crossed with 76-5EN pf2 mt-, the revertant (wild-type) phenotype segregated in a 2:2 pattern with the 76-5EN acetate-requiring phenotype, which indicates that the revertant results from a nuclear-gene mutation (Hong 1996). R107-4A likely results from true reversion or intragenic suppression within the 76-5EN gene.

Hong S (1996) Nuclear mutations affect the catalysis, stability, and expression of chloroplast ribulose-1,5-bisphosphate carboxylase/oxygenase in Chlamydomonas reinhardtii. Ph. D. thesis, University of Nebraska

Hong S, Spreitzer RJ (1994) Nuclear mutation inhibits expression of the chloroplast gene that encodes the large subunit of ribulose-1,5-bisphosphate carboxylase/oxygenase. Plant Physiol 106:673-678

Hong S, Spreitzer RJ (1998) Nuclear-gene mutations suppress a defect in the expression of the chloroplast-encoded large subunit of ribulosebisphosphate carboxylase/oxygenase. Plant Physiol 116:1387-1392

  • Locus:
  • PF2
  • Chromosome:
  • 11

From Robert J. Spreitzer, University of Nebraska, March 2014

Phenotype: requires acetate at 35 °C, temperature-conditional

This photosynthesis-competent revertant was selected from the Rubisco rbcL-mRNA deficient 76-5EN nuclear mutant (Hong and Spreitzer 1994, 1998). It can grow on minimal medium at 25 °C, but dies on minimal medium at 35 °C. R107-4B is a double mutant (76-5EN/S107-4B) that results from a nuclear-gene intergenic suppressor (S107-4B) of 76-5EN. When separated from 76-5EN, S107-4B has a temperature-conditional phenotype (Hong and Spreitzer 1998). This strain has been maintained with acetate medium in darkness since its isolation.

Hong S, Spreitzer RJ (1994) Nuclear mutation inhibits expression of the chloroplast gene that encodes the large subunit of ribulose-1,5-bisphosphate carboxylase/oxygenase. Plant Physiol 106:673-678

Hong S, Spreitzer RJ (1998) Nuclear-gene mutations suppress a defect in the expression of the chloroplast-encoded large subunit of ribulosebisphosphate carboxylase/oxygenase. Plant Physiol 116:1387-1392

From Robert J. Spreitzer, University of Nebraska, March 2014

Phenotype: requires acetate at 35 °C, temperature-conditional

Revertant R107-4B mt+ (76-5EN/S107-4B) was crossed with pf2 mt- to recover the nuclear intergenic suppressor S107-4B in the absence of 76-5EN. S107-4B alone does not affect the level of Rubisco rbcL mRNA, but it has a photosynthesis-deficient temperature-conditional phenotype (Hong and Spreitzer 1998). It can grow on minimal medium at 25 °C, but dies on minimal medium at 35 °C. This strain has been maintained with acetate medium in darkness since its isolation.

Hong S, Spreitzer RJ (1994) Nuclear mutation inhibits expression of the chloroplast gene that encodes the large subunit of ribulose-1,5-bisphosphate carboxylase/oxygenase. Plant Physiol 106:673-678

Hong S, Spreitzer RJ (1998) Nuclear-gene mutations suppress a defect in the expression of the chloroplast-encoded large subunit of ribulosebisphosphate carboxylase/oxygenase. Plant Physiol 116:1387-1392

  • Locus:
  • PF2
  • Chromosome:
  • 11

From Robert J. Spreitzer, University of Nebraska, March 2014

Phenotype: requires acetate at 35 °C, temperature-conditional

Revertant R107-4B mt+ (76-5EN/S107-4B) was crossed with pf2 mt- to recover the nuclear intergenic suppressor S107-4B in the absence of 76-5EN. S107-4B alone does not affect the level of Rubisco rbcL mRNA, but it has a photosynthesis-deficient temperature-conditional phenotype (Hong and Spreitzer 1998). It can grow on minimal medium at 25 °C, but dies on minimal medium at 35 °C. This strain has been maintained with acetate medium in darkness since its isolation.

Hong S, Spreitzer RJ (1994) Nuclear mutation inhibits expression of the chloroplast gene that encodes the large subunit of ribulose-1,5-bisphosphate carboxylase/oxygenase. Plant Physiol 106:673-678

Hong S, Spreitzer RJ (1998) Nuclear-gene mutations suppress a defect in the expression of the chloroplast-encoded large subunit of ribulosebisphosphate carboxylase/oxygenase. Plant Physiol 116:1387-1392