Table 1.

Reagents and resources

Reagent or resourceSourceIdentifier
Antibodies 
His Tag biotinylated antibody R&D Systems BAM050 
Biological samples   
Purified tubulin from bovine brains Schenk Packing Co. NA 
Yeast whole-cell lysate Listed strains NA 
Chemicals, peptides, and recombinant proteins 
Tubulin protein (fluorescent HiLyte 647): Porcine brain Cytoskeleton, Inc. TL670M 
Tubulin protein (biotin): Porcine brain Cytoskeleton, Inc, T333P 
Vectabond reagent, tissue section adhesion Vector Laboratories SP-1800-7 
mPEG-SVA MW 5,000 Laysan Bio, Inc. mPEG-succinimidyl valerate 
Biotinylated mPEG-SVA MW 5,000 Laysan Bio, Inc. Biotin-PEG-SVA 
Avidin DN Vector Laboratories A-3100-1 
Biotin-BSA Vector Laboratories B-2007-10 
Paclitaxel Millipore Sigma T7191-5MG 
Guanosine 5′-triphosphate sodium salt hydrate Milipore Sigma G8877-1G 
Streptavidin-coated polystyrene beads Spherotech SVP-05-10 
Guanosine-5′-[(α,β)-methyleno]triphosphate, sodium salt (GMPcPP) Jena Bioscience JBS-NU-405S 
Experimental models: Organisms/strains 
S. cerevisiae: MATa ura3-1::pCSE4-CSE4-XbaI(GFP):URA3 leu2,3-112 his3-11 trp1-1 ade2-1 LYS2+ can1-100 bar1 cse4Δ::KanMX Biggins lab SBY19926 
S. cerevisiae: MATa pDsn1-Dsn1-2D-3FLAG:URA3 Ndc10-GFP:kanMX6 Biggins lab SBY21618 
S. cerevisiae: MATa pDsn1-Dsn1-2D-3FLAG:URA3 Ndc80-GFP:kanMX6 Biggins lab SBY21620 
S. cerevisiae: MATa pDsn1-Dsn1-2D-3FLAG:URA3 Ctf19-GFP:kanMX6 Biggins lab SBY21621 
S. cerevisiae: MATx pDsn1-Dsn1-2D-3FLAG:URA3 Dam1-3GFP:HIS Biggins lab SBY20634 
Oligonucleotides 
Forward primer for generating 208-bp CEN3 or CEN3mut: /5BiosG/5′-GGTGGTT​CTGG​TGGTT​CTGG​TGAA​TTC-CCATT​CAATG​AAAT​ATATA​TTTC​TTACT​AT​TTC-3′ This paper 50_JDL 
Reverse primer for generating 208-bp CEN3 or CEN3mut: /5Atto565N/5′-GCT​ATT​CAT​TGA​AAAA​ATAG​TACA​AAT​AAG-3′ This paper 52_JDL 
5′ primer to tag Ndc80: 5′-ACG​AAA​TTT​GGA​GTT​TGA​AAC​TGA​ACA​TAA​CGT​AAC​AAA​TCG​GAT​CCC​CGG​GTT​AAT​TAA-3′ This paper 30_JDL 
3′ primer to tag Ndc80: 5′-CTG​TAG​ATT​GCT​CGG​GTA​TTA​TAT​ATC​ATT​TAT​TTT​ATT​AGA​ATT​CGA​GCT​CGT​TTA​AAC-3′ This paper 31_JDL 
5′ primer to tag Ctf19: 5′-GAT​CTG​CAA​CGT​TTG​CCT​ATT​CCC​GGA​CAT​GTA​CGC​CAG​GCG​GAT​CCC​CGG​GTT​AAT​TAA-3′ This paper 10_JDL 
3′ primer to tag Ctf19: 5′-TAA​GCA​AGC​CGT​CCA​GTT​GGC​AAT​GGC​AAA​TGG​AAC​ATC​AGA​ATT​CGA​GCT​CGT​TTA​AAC-3′ This paper 11_JDL 
5′ primer to tag Ndc10: 5′-TCA​AAA​TTC​ATT​TGA​TGG​TCT​GTT​AGT​ATA​TCT​ATC​TAA​CCG​GAT​CCC​CGG​GTT​AAT​TAA-3′ This paper 6_JDL 
3′ primer to tag Ndc10: 5′-TAT​CCC​TAT​ACG​AAA​CAG​TTT​AAA​CTT​CGA​AGC​TCC​CTC​AGA​ATT​CGA​GCT​CGT​TTA​AAC-3′ This paper 7_JDL 
Recombinant DNA 
pSB963: WT CEN3, 8LacO, TRP1 Lang et al. (2018)  NA 
pSB972: Mutant CEN3 (CCG->AGC CEN mutant), 8 LacO, TRP1 Lang et al. (2018)  NA 
Software and algorithms 
Single molecule colocalization analysis (written in LabView) This paper https://github.com/casbury69/smTIRF-spot-selection-colocalization-and-brightness-vs-time 
Displacement analysis (written in Igor Pro) This paper https://github.com/casbury69/flip-flop-IGOR-analysis-routines 
ImageJ (Fiji) Schindelin et al. (2012)  https://imagej.net/software/fiji/ 
MOSAICsuite Sbalzarini and Koumoutsakos (2005)  https://imagej.net/plugins/mosaicsuite 
Reagent or resourceSourceIdentifier
Antibodies 
His Tag biotinylated antibody R&D Systems BAM050 
Biological samples   
Purified tubulin from bovine brains Schenk Packing Co. NA 
Yeast whole-cell lysate Listed strains NA 
Chemicals, peptides, and recombinant proteins 
Tubulin protein (fluorescent HiLyte 647): Porcine brain Cytoskeleton, Inc. TL670M 
Tubulin protein (biotin): Porcine brain Cytoskeleton, Inc, T333P 
Vectabond reagent, tissue section adhesion Vector Laboratories SP-1800-7 
mPEG-SVA MW 5,000 Laysan Bio, Inc. mPEG-succinimidyl valerate 
Biotinylated mPEG-SVA MW 5,000 Laysan Bio, Inc. Biotin-PEG-SVA 
Avidin DN Vector Laboratories A-3100-1 
Biotin-BSA Vector Laboratories B-2007-10 
Paclitaxel Millipore Sigma T7191-5MG 
Guanosine 5′-triphosphate sodium salt hydrate Milipore Sigma G8877-1G 
Streptavidin-coated polystyrene beads Spherotech SVP-05-10 
Guanosine-5′-[(α,β)-methyleno]triphosphate, sodium salt (GMPcPP) Jena Bioscience JBS-NU-405S 
Experimental models: Organisms/strains 
S. cerevisiae: MATa ura3-1::pCSE4-CSE4-XbaI(GFP):URA3 leu2,3-112 his3-11 trp1-1 ade2-1 LYS2+ can1-100 bar1 cse4Δ::KanMX Biggins lab SBY19926 
S. cerevisiae: MATa pDsn1-Dsn1-2D-3FLAG:URA3 Ndc10-GFP:kanMX6 Biggins lab SBY21618 
S. cerevisiae: MATa pDsn1-Dsn1-2D-3FLAG:URA3 Ndc80-GFP:kanMX6 Biggins lab SBY21620 
S. cerevisiae: MATa pDsn1-Dsn1-2D-3FLAG:URA3 Ctf19-GFP:kanMX6 Biggins lab SBY21621 
S. cerevisiae: MATx pDsn1-Dsn1-2D-3FLAG:URA3 Dam1-3GFP:HIS Biggins lab SBY20634 
Oligonucleotides 
Forward primer for generating 208-bp CEN3 or CEN3mut: /5BiosG/5′-GGTGGTT​CTGG​TGGTT​CTGG​TGAA​TTC-CCATT​CAATG​AAAT​ATATA​TTTC​TTACT​AT​TTC-3′ This paper 50_JDL 
Reverse primer for generating 208-bp CEN3 or CEN3mut: /5Atto565N/5′-GCT​ATT​CAT​TGA​AAAA​ATAG​TACA​AAT​AAG-3′ This paper 52_JDL 
5′ primer to tag Ndc80: 5′-ACG​AAA​TTT​GGA​GTT​TGA​AAC​TGA​ACA​TAA​CGT​AAC​AAA​TCG​GAT​CCC​CGG​GTT​AAT​TAA-3′ This paper 30_JDL 
3′ primer to tag Ndc80: 5′-CTG​TAG​ATT​GCT​CGG​GTA​TTA​TAT​ATC​ATT​TAT​TTT​ATT​AGA​ATT​CGA​GCT​CGT​TTA​AAC-3′ This paper 31_JDL 
5′ primer to tag Ctf19: 5′-GAT​CTG​CAA​CGT​TTG​CCT​ATT​CCC​GGA​CAT​GTA​CGC​CAG​GCG​GAT​CCC​CGG​GTT​AAT​TAA-3′ This paper 10_JDL 
3′ primer to tag Ctf19: 5′-TAA​GCA​AGC​CGT​CCA​GTT​GGC​AAT​GGC​AAA​TGG​AAC​ATC​AGA​ATT​CGA​GCT​CGT​TTA​AAC-3′ This paper 11_JDL 
5′ primer to tag Ndc10: 5′-TCA​AAA​TTC​ATT​TGA​TGG​TCT​GTT​AGT​ATA​TCT​ATC​TAA​CCG​GAT​CCC​CGG​GTT​AAT​TAA-3′ This paper 6_JDL 
3′ primer to tag Ndc10: 5′-TAT​CCC​TAT​ACG​AAA​CAG​TTT​AAA​CTT​CGA​AGC​TCC​CTC​AGA​ATT​CGA​GCT​CGT​TTA​AAC-3′ This paper 7_JDL 
Recombinant DNA 
pSB963: WT CEN3, 8LacO, TRP1 Lang et al. (2018)  NA 
pSB972: Mutant CEN3 (CCG->AGC CEN mutant), 8 LacO, TRP1 Lang et al. (2018)  NA 
Software and algorithms 
Single molecule colocalization analysis (written in LabView) This paper https://github.com/casbury69/smTIRF-spot-selection-colocalization-and-brightness-vs-time 
Displacement analysis (written in Igor Pro) This paper https://github.com/casbury69/flip-flop-IGOR-analysis-routines 
ImageJ (Fiji) Schindelin et al. (2012)  https://imagej.net/software/fiji/ 
MOSAICsuite Sbalzarini and Koumoutsakos (2005)  https://imagej.net/plugins/mosaicsuite 
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