Table II.
S. cerevisiae strains used in this study
| Strain | Genotype | Reference or source |
| YEF473A | a his3 leu2 lys2 trp1 ura3 | (Bi and Pringle, 1996) |
| YEF1820 | a/α his3/his3 leu2/leu2 lys2/lys2 trp1/trp1 ura3/ura3 myo1Δ::KanMX6/myo1Δ::HIS3 | (Bi et al., 1998) |
| YEF1804 | As YEF473A except myo1Δ::KanMX6 | (Bi et al., 1998) |
| YEF5762 | As YEF473A except CHS2-GFP:His3MX6 | This studya |
| YEF5804 | As YEF473A except CDC3-mCherry:LEU2 | This studyb |
| YEF5874 | As YEF473A except CHS2-GFP:His3MX6 CDC3-mCherry:LEU2 | This studyc |
| YEF5986 | As YEF473A except MYO2-GFP:His3 CDC3-mCherry:LEU2 | This studyc |
| YEF6179 | a mlc1-11 MYO1-GFP:KanMX6 CDC3-mCherry:LEU2 | This studyd |
| YEF6113 | a MYO1-GFP:His3MX6 NUP57-GFP:URA3 his3 leu2 lys2 ura3 | This studye |
| YEF6307 | a MYO1-GFP NUP57-GFP:URA3 | This studyf |
| YEF6308 | a myo1-TD2Δ-GFP NUP57-GFP:URA3 | This studyf |
| YEF6309 | a myo1-Tail-GFP NUP57-GFP:URA3 | This studyf |
| YEF6310 | a myo1-TD2-GFP NUP57-GFP:URA3 | This studyf |
| YEF6316 | a bni5Δ::His3MX6 MYO1-GFP NUP57-GFP:URA3 | This studyf |
| YEF6321 | a MYO1-GFP BNI5-mCherry:KanMX6 | This studyg |
| YEF6322 | a bni5Δ::His3MX6 myo1-TD2Δ-GFP | This studyh |
| YEF6323 | a bni5Δ::His3MX6 myo1-TD2Δ-GFP CDC3-mCherry:LEU2 | This studyh |
| YEF6325 | a iqg1Δ::His3MX6 CDC3-mCherry:LEU2 [pRS316-MYO1-C-GFP] | This studyi |
| YEF6326 | a myo1-mTD1-GFP BNI5-mCherry:KanMX6 | This studyj |
| Y5005-8D | As YEF473A except mlc1-11 | (Luo et al., 2004) |
| YKT520 | As YEF473A except MYO2-GFP:HIS3 | K. Tanaka |
| YKT662 | As YEF473A except MYO2-GFP:TRP1 | K. Tanaka |
| RNY970 | As YEF473A except iqg1Δ::His3MX6 [YCp50-IQG1] | J.R. Pringle |
| RNY2594 | As YEF473A except iqg1Δ::His3MX6 MYO1-GFP:KanMX6 [pRS315-IQG1-GST] | This studyk |
| RNY2595 | As YEF473A except iqg1Δ::His3MX6 MYO1-GFP:KanMX6 [pRS315-IQG1(Δ2-411)-GST] | This studyk |
| RNY2596 | As YEF473A except iqg1Δ::His3MX6 MYO1-GFP:KanMX6 CDC3-mCherry:URA3 [pRS315-IQG1-GST] | This studyl |
| RNY2597 | As YEF473A except iqg1Δ::His3MX6 MYO1-GFP:KanMX6 CDC3-mCherry:URA3 [pRS315-IQG1(Δ2-411)-GST] | This studyl |
| Masa1243 | As YEF473A except myo1Δ::URA3-KanMX6 [pUG23-MYO1] | See text |
| YJL221A | As YEF473A except myo1-TD1-GFP | See text |
| YJL222A | As YEF473A except myo1-TD2-GFP | See text |
| YJL335A | As YEF473A except myo1-Tail-GFP | See text |
| YJL376A | As YEF473A except myo1-AD-GFP | See text |
| YJL488A | As YEF473A except myo1-TD2Δ-GFP | See text |
| YJL489A | As YEF473A except myo1-mTD1-GFP | See text |
| XDY7 | YEF1820 [pRS316-MYO1-C-TAP] | This study |
| XDY41 | As YEF473A except MYO1-GFP | See text |
| XDY99 | YEF1820 [pRS316-MYO1-TD2Δ-C-TAP] | This study |
| XDY154 | YEF473A [pRS316-MYO1-TD2-GFP] | This studym |
| XDY173 | a mlc1-11 [pRS316-MYO1-TD2-GFP] | This studym |
| XDY189 | a myo1-mTD1-GFP BNI5-3HA:His3MX6 | This studyn |
| XDY190 | a myo1-Tail-GFP BNI5-3HA:His3MX6 | This studyn |
| XDY191 | a myo1-TD1-GFP BNI5-3HA:His3MX6 | This studyn |
| XDY192 | a myo1-TD2-GFP BNI5-3HA:His3MX6 | This studyn |
| XDY194 | a myo1-AD-GFP BNI5-3HA:His3MX6 | This studyn |
| XDY211 | a myo1-mTD1-GFP BNI5-mCherry:KanMX6 [pRS316-N-NotI-MYO1] | This studyo |
| XDY218 | a iqg1Δ::His3MX6 [pRS316-MYO1-TD2-GFP] | This studyp |
| XDY254 | a MYO1-GFP bni5Δ::His3MX6 | This studyn |
| XDY257 | a myo1-TD2-GFP bni5Δ::His3MX6 | This studyn |
| XDY258 | a myo1-mTD1-GFP bni5Δ::His3MX6 | This studyn |
| XDY286 | a MYO1-GFP CDC3-mCherry:LEU2 | This studyq |
| XDY287 | a bni5Δ::His3MX6 MYO1-GFP CDC3-mCherry:LEU2 | This studyq |
| XDY288 | a myo1-Tail-GFP CDC3-mCherry:LEU2 | This studyq |
| XDY289 | a myo1-TD2Δ-GFP CDC3-mCherry:LEU2 | This studyq |
| XDY290 | a myo1-TD2-GFP CDC3-mCherry:LEU2 | This studyq |
| XDY295 | a iqg1Δ::His3MX6 [pRS316-MYO1-TD2-GFP] [pRS315-IQG1-GST] | This studyp |
| XDY296 | a iqg1Δ::His3MX6 [pRS316-MYO1-TD2-GFP] [pRS315-IQG1(1-697)-GST] | This studyp |
| XDY300 | a iqg1Δ::His3MX6 [pRS316-MYO1-TD2-GFP] [pRS315-IQG1(Δ2-411)-GST] | This studyp |
Constructed by a PCR-based method (Longtine et al., 1998).
Constructed by transforming BglII-digested YIp128-CDC3-mCherry (Gao et al., 2007) into YEF473A and integrating at the CDC3 locus.
Constructed by PCR-amplifying CHS2-GFP:HIS3MX6 and MYO2-GFP:HIS3 fragments from YEF5762 and YKT520, respectively, and transforming into YEF5804.
Constructed by transforming Y5005-8D with a MYO1-GFP:KanMX6 fragment that was PCR-amplified from YEF2293 (Luo et al., 2004), followed by integration of BglII-digested YIp128-CDC3-mCherry at the CDC3 locus.
Constructed by a PCR-based method(Longtine et al., 1998) using plasmids pFA6a-GFP-His3MX6 (Longtine et al., 1998) and pFA6a-GFP-URA3 (provided by C. Burd, University of Pennsylvania, Philadelphia, PA) as templates for tagging MYO1 and NUP57, respectively. The PCR fragments containing MYO1-GFP:His3MX6 and NUP57-GFP:URA3 were transformed sequentially into strain RLY1 (Winter et al., 1999).
Constructed by transforming PCR-amplified NUP57-GFP:URA3 from YEF6113 into strains XDY41, YJL488A, YJL335A, YJL222A, and XDY254, respectively.
Constructed by transforming PCR-amplified BNI5-mCherry:KanMX6 from XDY211 into XDY41.
Constructed by transforming PCR-amplified bni5Δ::His3MX6 from XDY254 into YJL448A and XDY289, respectively.
Strain XDY218 was first transformed by BglII-digested YIp128-CDC3-mCherry and then streaked on a SC-Leu + 5-FOA plate to select for the loss of plasmid pRS316-MYO1-TD2-GFP. The resulting strain was then transformed with plasmid pRS316-MYO1-C-GFP.
Constructed by streaking XDY211 on a SC + 5-FOA plate to select for the loss of plasmid pRS316-N-NotI-MYO1.
Strain RNY1264 (iqg1Δ::His3MX6 MYO1-GFP:KanMX6 [YCp50-IQG1]) was transformed with pRS315-IQG1-GST or pRS315-IQG1(Δ2-411)-GST, and the resulting strains were streaked on a SC-Leu + 5-FOA plate to eliminate YCp50-IQG1.
Constructed by transforming BglII-digested YIp211-CDC3-mCherry into strains RNY2594 and RNY2595 and integrating at the CDC3 locus.
Constructed by transforming the plasmid into YEF473A and Y5005-8D, respectively.
Strains carrying different myo1 alleles were transformed with plasmid pRS316-MYO1. The resulting strains were either 3HA-tagged or deleted for BNI5 using a PCR-based method (Longtine et al., 1998), and then selected for the loss of the MYO1 plasmid by growing cells in non-selective YPD medium.
Constructed by transforming a BNI5-mCherry:KanMX6 fragment that was PCR-amplified from plasmid pFA6a-mCherry-KanMX6 into YJL489A carrying plasmid pRS316-N-NotI-MYO1 (Caviston et al., 2003).
Strain RNY970 (shown in the table) was streaked from a SC-His plate onto a SC-His + 5-FOA plate; the resulting single colonies were checked by PCR to confirm the loss of the IQG1 plasmid. The resulting iqg1Δ::His3MX6 strain was then transformed with indicated plasmids to generate strains XDY218, XDY295, XDY296, and XDY300.
Strains XDY41, XDY287, YJL335A, YJY222A, and YJY488A were transformed with plasmid pRS316-MYO1. The resulting strains were then transformed with BglII-digested integrative plasmid YIp128-CDC3-mCherry and transformants were selected on SC-Leu plates at 25°C and then grown in non-selective YPD medium to lose the MYO1 plasmid.