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pYES2 vector (V010202)

Price Information

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V010202 pYES2 In stock, instant shipping

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Two tubes of lyophilized plasmid will be delivered, each tube is about 5µg.

Basic Vector Information

The pYES2 vector is an episomal high-copy plasmid designed for galactose-inducible protein expression in Saccharomyces cerevisiae.

Vector Name:
pYES2
Antibiotic Resistance:
Ampicillin
Length:
5860 bp
Type:
Yeast Plasmids
Replication origin:
ori
Source/Author:
Invitrogen (Life Technologies)
Copy Number:
High copy number
Promoter:
GAL1
Growth Strain(s):
stbl3
Growth Temperature:
37℃

pYES2 vector Map

Copy Sequence View Map
pYES25860 bp60012001800240030003600420048005400GAL1 promoterT7 promoterMCSCYC1 terminatororiAmpRURA3URA3 promoter2u oriM13 ori

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

  • Tsai HC, Hsieh CH, Hsu CW, Hsu YH, Chien LF. Cloning and Organelle Expression of Bamboo Mitochondrial Complex I Subunits Nad1, Nad2, Nad4, and Nad5 in the Yeast Saccharomyces cerevisiae. Int J Mol Sci. 2022 Apr 6;23(7):4054. doi: 10.3390/ijms23074054. PMID: 35409414; PMCID: PMC8999482.

  • Lin CC, Hsieh MH, Teng SC. Genistein suppresses the proliferation of telomerase-negative cells. Food Sci Nutr. 2016 May 26;5(2):197-204. doi: 10.1002/fsn3.382. PMID: 28265354; PMCID: PMC5332266.

  • Piraine REA, Gonçalves VS, Dos Santos Junior AG, Cunha RC, de Albuquerque PMM, Conrad NL, Leite FPL. Expression cassette and plasmid construction for Yeast Surface Display in Saccharomyces cerevisiae. Biotechnol Lett. 2021 Aug;43(8):1649-1657. doi: 10.1007/s10529-021-03142-w. Epub 2021 May 2. PMID: 33934257.

pYES2 vector Sequence

LOCUS       Exported                5860 bp DNA     circular SYN 02-SEP-2024
DEFINITION  High-copy episomal vector for galactose-inducible expression of 
            proteins in Saccharomyces cerevisiae.
ACCESSION   .
VERSION     .
KEYWORDS    pYES2
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5860)
  AUTHORS   Invitrogen (Life Technologies)
  TITLE     Direct Submission
REFERENCE   2  (bases 1 to 5860)
  TITLE     Direct Submission
REFERENCE   3  (bases 1 to 5860)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"
COMMENT     SGRef: number: 2; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..5860
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     source          1..5858
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        2..443
                     /label=GAL1 promoter
                     /note="inducible promoter, regulated by Gal4"
     promoter        475..493
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     misc_feature    501..600
                     /label=MCS
                     /note="MCS"
                     /note="multiple cloning site"
     terminator      609..856
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     rep_origin      complement(1104..1692)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(1866..2723)
                     /codon_start=1
                     /label=AmpR
                     /note="beta-lactamase"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     CDS             complement(2822..3622)
                     /label=URA3
                     /note="orotidine-5'-phosphate decarboxylase, required for
                     uracil biosynthesis"
     promoter        complement(3623..3838)
                     /label=URA3 promoter
     rep_origin      complement(4439..5316)
                     /direction=LEFT
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
     rep_origin      complement(5327..5840)
                     /direction=LEFT
                     /label=M13 ori
                     /note="M13 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"