WormBase Tree Display for Construct: WBCnstr00001895
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| WBCnstr00001895 | Public_name | pJK909 | |
|---|---|---|---|
| Summary | [pop-1p::POP-1::GFP] | ||
| Driven_by_gene | WBGene00004077 | ||
| Gene | WBGene00004077 | ||
| Fusion_reporter | GFP | ||
| Type_of_construct | Cterminal_translational_fusion | ||
| Construction_summary | The POP-1::GFP construct included 3.5 kb of sequence upstream of the pop-1 gene, introns 1 and 2, and GFP fused at the C terminus of the protein; this construct was produced by a PCR ligation technique. The pop-1 cDNA::GFP fusion was made by digestion of pJK706 with BbsI, end filling with Klenow, and then digesting with HindIII to remove the pop-1 fragment. The GFP vector pPD95.79 was digested with HindIII and SmaI and ligated with the HindIII BbsI(blunt) fragment from pJK706 to produce plasmid pJK909. The following fragments were then produced either by PCR using the Expand 20KbPLUS system (Roche) or by digestion and gel purification: (1) a genomic fragment consisting of 3.5 kb upstream of the start codon and continuing through exon 2 (primer sequences: 5-AGCAAGGTGTCTACTGTCG CCTGTC-3 and 5-TTTTCGCCAATTTTTATGTGT-3), (2) a genomic fragment containing exon 1 and continuing into exon 3 (primer sequences: 5-ATGGCCTAACTTCCGC-3 and 5-TTTCGCCTGTTCTTCCTTCGA-3), and (3) a PvuI fragment of pJK909 that begins in exon 3 of the pop-1 cDNA::GFP fusion and continues through the unc-54 3-UTR from pPD95.79. These three fragments were produced in duplicate and all were combined and used as the template in PCR reactions to amplify the entire POP-1::GFP product using the Expand 20KbPLUS system (Roche; primer sequences 5-AGC AAGGTGTCTACTGTCGCCTGTC-3 and 5-GAGGTTTTCA CCGTCATCACC-3). | ||
| Used_for | Transgene_construct | WBTransgene00027280 | |
| Reference | WBPaper00006440 |
