p5E-nos2 promoter_L4-R1
(Plasmid #226796)

• Purpose: 5'-entry vector with the nanos2 promoter cloned from Danio rerio. For use in Gateway cloning. Includes a red heart (pBH; cmlc2:RFP) marker.
• Depositing Lab: Florence Marlow
• Publication: Primordial germ cells differentially contribute to the germline in zebrafish. (unpublished)

Backbone

• Vector backbone: p5E-MCS
• Backbone manufacturer: Nathan Lawson
• Backbone size w/o insert (bp): 2810
• Total vector size (bp): 5238
• Vector type: Bacterial Expression

Growth in Bacteria

• Bacterial Resistance(s): Kanamycin, 50 μg/mL
• Growth Temperature: 37°C
• Growth Strain(s): DH5alpha
• Copy number: Unknown

Gene/Insert

• Gene/Insert name: nanos2
• Species: D. rerio (zebrafish)
• Insert Size (bp): 2417
• Entrez Gene: nanos2

Cloning Information

• Cloning method: Restriction Enzyme
• 5′ cloning site: SmaI (destroyed during cloning)
• 3′ cloning site: None (unknown if destroyed)
• 5′ sequencing primer: M13F

Terms and Licenses

• Academic/Nonprofit Terms:
  • UBMTA
• Industry Terms:
  • Not Available to Industry

Depositor Comments

Cloned by Maya Pahima.
The putative zebrafish nos2 promoter was cloned from zebrafish ovary gDNA from a region directly upstream of the nos2 coding sequence with the following primers: 3’-ACTGGCCTAAAGGGTGTCCT-5’ and 3’-CGTCCCTTGCCTTTAGTCTG-5’. The cloned amplicon was inserted into the pCR8 cloning vector using the TOPO™ TA Cloning Kit (Thermo Fisher, K250020) for stabilization and sequencing. Putative nos2 promoter region was excised from the TOPO vector using EcoRI and SmaI, treated with T4 polymerase, then inserted into SmaI-linearized p5E-MCS (Addgene, #26029).