Price Information
| Cat No. | Plasmid Name | Availability | Add to cart |
|---|---|---|---|
| V010194 | YCplac111 | In stock, instant shipping |
Buy one, get one free! |
Two tubes of lyophilized plasmid will be delivered, each tube is about 5µg.
Basic Vector Information
YCplac111 is a plasmid with ARS1, CEN4, and LEU2. It's used to study DNA replication and maintenance in yeast and to measure the ability of cells to maintain minichromosomes.
- Vector Name:
- YCplac111
- Antibiotic Resistance:
- Ampicillin
- Length:
- 6103 bp
- Type:
- Yeast Plasmids
- Replication origin:
- ori
- Host:
- Yeast
- Source/Author:
- Gietz RD, Sugino A.
- Selection Marker:
- LEU2
- Copy Number:
- High copy number
- Promoter:
- LEU2
- 5' Primer:
- M13 fwd
- 3' Primer:
- M13 rev
- Growth Strain(s):
- DH10B
- Growth Temperature:
- 37℃
YCplac111 vector Map
Plasmid Protocol
1. Centrifuge at 5,000×g for 5 min.
2. Carefully open the tube and add 20 μl of sterile water to dissolve the DNA.
3. Close the tube and incubate for 10 minutes at room temperature.
4. Briefly vortex the tube and then do a quick spin to concentrate the liquid at the bottom. Speed is less than 5000×g.
5. Store the plasmid at -20 ℃.
6. The concentration of plasmid re-measurement sometimes differs from the nominal value, which may be due to the position of the lyophilized plasmid in the tube, the efficiency of the re-dissolution, the measurement bias, and adsorption on the wall of the tube, therefore, it is recommended to transform and extract the plasmid before using it
General Plasmid Transform Protocol
1. Take one 100μl of the competent cells and thaw it on ice for 10min, add 2μl of plasmid, then ice bath for 30min, then heat-shock it at 42℃ for 60s, do not stir, and then ice bath for 2min.
2. Add 900μl of LB liquid medium without antibiotics, and incubate at 37℃ for 45min (30℃ for 1-1.5 hours) with 180rpm shaking.
3. Centrifuge at 6000rpm for 5min, leave only 100μl of supernatant to resuspend the bacterial precipitate and spread it onto the target plasmid-resistant LB plate.
4. Invert the plate and incubate at 37℃ for 14h, or at 30℃ for 20h.
5. Pick a single colony into LB liquid medium, add the corresponding antibiotics, incubate at 220rpm for 14h, and extract the plasmid according to the experimental needs and the instructions of the plasmid extraction kit.
References
- Rizzardi LF, Dorn ES, Strahl BD, Cook JG. DNA replication origin function is promoted by H3K4 di-methylation in Saccharomyces cerevisiae. Genetics. 2012 Oct;192(2):371-84. doi: 10.1534/genetics.112.142349. Epub 2012 Jul 30. PMID: 22851644; PMCID: PMC3454870.
YCplac111 vector Sequence
LOCUS 40924_49462 6103 bp DNA circular SYN 01-JAN-1980
DEFINITION Yeast centromeric plasmid with a LEU2 marker.
ACCESSION .
VERSION .
KEYWORDS .
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 6103)
AUTHORS Gietz RD, Sugino A.
TITLE New yeast-Escherichia coli shuttle vectors constructed with in vitro
mutagenized yeast genes lacking six-base pair restriction sites.
JOURNAL Gene 1988;74:527-34.
PUBMED 3073106
REFERENCE 2 (bases 1 to 6103)
TITLE Direct Submission
REFERENCE 3 (bases 1 to 6103)
AUTHORS .
TITLE Direct Submission
COMMENT SGRef: number: 1; type: "Journal Article"; journalName: "Gene";
date: "1988"; volume: "74"; pages: "527-34"
COMMENT SGRef: number: 2; type: "Journal Article"
COMMENT The sequence of this plasmid was reconstructed using the description
from Gietz and Sugino.
FEATURES Location/Qualifiers
source 1..6103
/mol_type="other DNA"
/organism="synthetic DNA construct"
protein_bind 106..127
/label=CAP binding site
/note="CAP binding activates transcription in the presence
of cAMP."
promoter 142..172
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 180..196
/label=lac operator
/note="The lac repressor binds to the lac operator to
inhibit transcription in E. coli. This inhibition can be
relieved by adding lactose or
isopropyl-beta-D-thiogalactopyranoside (IPTG)."
primer_bind 204..220
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
misc_feature complement(233..289)
/label=MCS
/note="pUC18/19 multiple cloning site"
primer_bind complement(290..306)
/label=M13 fwd
/note="common sequencing primer, one of multiple similar
variants"
misc_feature complement(1064..2227)
/label=CEN/ARS
/note="S. cerevisiae CEN4 centromere fused to the
autonomously replicating sequence ARS1/ARS416"
CDS complement(2592..3683)
/codon_start=1
/label=LEU2
/note="3-isopropylmalate dehydrogenase, required for
leucine biosynthesis"
/translation="MSAPKKIVVLPGDHVGQEITAEAIKVLKAISDVRSNVKFDFENHL
IGGAAIDATGVPLPDEALEASKKVDAVLLGAVGGPKWGTGSVRPEQGLLKIRKELQLYA
NLRPCNFASDSLLDLSPIKPQFAKGTDFVVVRELVGGIYFGKRKEDDGDGVAWDSEQYT
VPEVQRITRMAAFMALQHEPPLPIWSLDKANVLASSRLWRKTVEETIKNEFPTLKVQHQ
LIDSAAMILVKNPTHLNGIIITSNMFGDIISDEASVIPGSLGLLPSASLASLPDKNTAF
GLYEPCHGSAPDLPKNKVNPIATILSAAMMLKLSLNLPEEGKAIEDAVKKVLDAGIRTG
DLGGSNSTTEVGDAVAEEVKKILA"
promoter complement(3684..4091)
/label=LEU2 promoter
promoter 4197..4301
/label=AmpR promoter
CDS 4302..5159
/codon_start=1
/label=AmpR
/note="beta-lactamase"
/translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
LIKHW"
rep_origin 5333..5921
/direction=RIGHT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"