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pNKY1009

87624

Clone type
Vector
Applications
Molecular biology
Product format
Frozen
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Price: $620.00 EA
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Documentation

ATCC determines the biosafety level of a material based on our risk assessment as guided by the current edition of Biosafety in Microbiological and Biomedical Laboratories (BMBL), U.S. Department of Health and Human Services. It is your responsibility to understand the hazards associated with the material per your organization’s policies and procedures as well as any other applicable regulations as enforced by your local or national agencies.

Detailed product information

General

Specific applications
contains sequence ATP phosphoribosyltransferase
contains sequence phosphoribosylanthranilate isomerase
marker deletion vector phosphoribosylanthranilate isomerase
produces protein uridine monophosphate synthetase UMP synthase, orotate phosphoribosyltransferase, orotidine 5'-phosphate decarboxylase, orotate phosphoribosyltransferase 1

Characteristics

Comments
Restriction digests of the clone give the following sizes (kb): BglII--9.6; EcoRI--5.2, 4.4; BglII/EcoRI--4.6, 4.4, 0.6.
The two step selection process requires a ura3 transformation host (this host can be created using pJL164 (ATCC 87471)). After transformation with the EcoRI/BglII digested plasmid, URA3 integrants are selected on ura- plates.
The deletion strain is then recovered by selection on 5-FOA plates (loss of URA3 marker by a homologous recombination event between the two hisG repeats).
E. coli containing this plasmid should be grown on media lacking pyrimidines to select for URA3-containing cells.
This deleter vector is used to create yeast strains with a trp1 auxotrophic marker deletion.
The 4.6 kb EcoRI/BglII insert contains two direct repeats of the Salmonella hisG gene flanking URA3 plus TRP1 sequences flanking the hisG-URA3-hisG sequence.
The plasmid was constructed by inserting the 3.8 kb BamHI-BglII hisG-URA3-hisG fragment into the modified EcoRV site within the TRP1 gene of YEp7.
Mycoplasma contamination
Not detected

Vector information

Construct size (kb)
9.60
Intact vector size
9.600
Vector name
pNKY1009 (plasmid)
Type of vector
plasmid
Construction
YRp7, pNKY51
Host range
Saccharomyces cerevisiae
Candida robusta
Escherichia coli
Vector information
Other unique sites: PvuII
Features (with orientation and position when available):
Coding sequence
3' TRP1, <-; hisG, ->; hisG, ->; 5' TRP1, <-; ROP, ->; 3' TRP1; 5' TRP1; ROP; hisG
Markers
ampR; URA3
Replicon
pMB1; ARS1, →
Restriction sites
BglII; EcoRI

Insert information

Target gene
ATP phosphoribosyltransferase; uridine monophosphate synthetase

Handling information

Host
Escherichia coli FD 27747 [DB6507] (ATCC 35673)
Medium
Temperature
37°C
Handling notes
Restriction digests of the clone give the following sizes (kb): BglII--9.6;
EcoRI--5.2, 4.4; BglII/EcoRI--4.6, 4.4, 0.6.
- ATCC staff

E. coli containing plasmid should be grown on medium lacking pyrimidines to
select for URA3-containing cells.
- personal communication

The 4.6 kb EcoRI/BglII insert contains two direct repeats of the Salmonella hisG
gene flanking URA3 plus TRP1 sequences flanking the hisG-URA3-hisG sequence.
- Genetics 116: 541-545, 1987

This deleter vector is used to create yeast strains with a trp1 auxotrophic
marker deletion.
- Genetics 116: 541-545, 1987

The two step selection process requires a ura3 transformation host (this host
can be created using pJL164 (ATCC 87471)). After transformation with the
EcoRI/BglII digested plasmid, URA3 integrants are selected on ura- plates.
- Genetics 116: 541-545, 1987

The deletion strain is then recovered by selection on 5-FOA plates (loss of URA3
marker by a homologous recombination event between the two hisG repeats).
- Genetics 116: 541-545, 1987

The plasmid was constructed by inserting the 3.8 kb BamHI-BglII hisG-URA3-hisG
fragment into the modified EcoRV site within the TRP1 gene of YEp7.
- Genetics 116: 541-545, 1987

History

Depositors
E Alani
Special collection
NCRR Contract

Legal disclaimers

Intended use
This product is intended for laboratory research use only. It is not intended for any animal or human therapeutic use, any human or animal consumption, or any diagnostic use.
Warranty

The product is provided 'AS IS' and the viability of ATCC® products is warranted for 30 days from the date of shipment, provided that the customer has stored and handled the product according to the information included on the product information sheet, website, and Certificate of Analysis. For living cultures, ATCC lists the media formulation and reagents that have been found to be effective for the product. While other unspecified media and reagents may also produce satisfactory results, a change in the ATCC and/or depositor-recommended protocols may affect the recovery, growth, and/or function of the product. If an alternative medium formulation or reagent is used, the ATCC warranty for viability is no longer valid.  Except as expressly set forth herein, no other warranties of any kind are provided, express or implied, including, but not limited to, any implied warranties of merchantability, fitness for a particular purpose, manufacture according to cGMP standards, typicality, safety, accuracy, and/or noninfringement.

Disclaimers

This product is intended for laboratory research use only. It is not intended for any animal or human therapeutic use, any human or animal consumption, or any diagnostic use. Any proposed commercial use is prohibited without a license from ATCC.

While ATCC uses reasonable efforts to include accurate and up-to-date information on this product sheet, ATCC makes no warranties or representations as to its accuracy. Citations from scientific literature and patents are provided for informational purposes only. ATCC does not warrant that such information has been confirmed to be accurate or complete and the customer bears the sole responsibility of confirming the accuracy and completeness of any such information.

This product is sent on the condition that the customer is responsible for and assumes all risk and responsibility in connection with the receipt, handling, storage, disposal, and use of the ATCC product including without limitation taking all appropriate safety and handling precautions to minimize health or environmental risk. As a condition of receiving the material, the customer agrees that any activity undertaken with the ATCC product and any progeny or modifications will be conducted in compliance with all applicable laws, regulations, and guidelines. This product is provided 'AS IS' with no representations or warranties whatsoever except as expressly set forth herein and in no event shall ATCC, its parents, subsidiaries, directors, officers, agents, employees, assigns, successors, and affiliates be liable for indirect, special, incidental, or consequential damages of any kind in connection with or arising out of the customer's use of the product. While reasonable effort is made to ensure authenticity and reliability of materials on deposit, ATCC is not liable for damages arising from the misidentification or misrepresentation of such materials.

Please see the material transfer agreement (MTA) for further details regarding the use of this product. The MTA is available at www.atcc.org.

Permits & Restrictions

Import Permit for the State of Hawaii

If shipping to the U.S. state of Hawaii, you must provide either an import permit or documentation stating that an import permit is not required. We cannot ship this item until we receive this documentation. Contact the Hawaii Department of Agriculture (HDOA), Plant Industry Division, Plant Quarantine Branch to determine if an import permit is required.

MORE INFORMATION ABOUT PERMITS AND RESTRICTIONS

Frequently Asked Questions

References

Curated Citations

Alani E, et al. A method for gene disruption that allows repeated use of URA3 selection in the construction of multiply disrupted yeast strains. Genetics 116: 541-545, 1987. PubMed: 3305158

Jef D Boeke, personal communication

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