WormBase Tree Display for Expr_pattern: Expr956
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| Expr956 | Expression_of | Gene | WBGene00003052 | |
|---|---|---|---|---|
| Reflects_endogenous_expression_of | WBGene00003052 | |||
| Homol | Homol_homol | DY3:Expr | ||
| Expression_data | Life_stage | WBls:0000002 | ||
| Anatomy_term | WBbt:0004017 | Certain | ||
| GO_term | GO:0034399 | |||
| Subcellular_localization | nuclear periphery of all cells | |||
| Type | Reporter_gene | Three different lmn-1-GFP constructs were used. The construct pDRNL1, contains a GFP-Ce-lamin fusion with GFP fused to the N-terminus of lmn-1: a 6.5-kb HindIISalI genomic fragment positioned between 7.95 kb and 1.45 kb upstream to the lmn-1 coding region, was cloned into the pPD95.77 vector. A 1.45-kb fragment, just 5' to the lmn-1 coding region, was amplified by PCR with a novel NcoI cloning site introduced at its 3' end, and inserted 3' to the 6.5-kb genomic fragment. The 0.75-kb KpnISmaI fragment of the GFP gene was PCR amplified from pEGFP and inserted into the NcoI and SmaI sites. A 2.2-kb KpnISmaI fragment of the lmn-1 genomic region, containing the complete lamin coding region, was PCR amplified and inserted 3' in frame to the GFP gene. The second construct is a fusion of GFP to the C-terminus of Ce-lamin. To prepare this construct, termed pJKL380.4, long-range PCR was used to amplify from wild-type genomic DNA a 9.6-kb lamin genomic fragment, containing 4.7 kb of 5' sequence, the entire coding sequence plus introns, and 2.8 kb of 3' sequence. This 9.6-kb fragment was engineered such that unique NotI and SmaI sites were present at its ends and used for cloning into the pBluescript II SK vector. The resulting plasmid, in which the BamHI site in the vector backbone was destroyed, contained a single BamHI site 12 amino acids before the stop codon of lamin. This site was used to insert GFP (from pPD102.33) in frame into the lmn-1 gene. A third construct, termed pDRCL2, contained 7.95 kb of 5' promoter sequence of lmn-1, the first three exons, and two introns of lmn-1 fused in frame to the GFP gene. --precise ends. | ||
| Pattern | With the exception of mature sperm, Ce-lamin-GFP fusion protein was localized to the nuclear periphery of all cells throughout the development of the nematode. The Ce-lamin-GFP expression in these lines reproduced the antibody staining pattern of the native protein, including the presence of subregions in the nuclear envelope with higher fluorescence intensity. | |||
| Picture | WBPicture0000008682 | |||
| Remark | data derived from more than one GFP construct. | |||
| Reference | WBPaper00004416 | |||
| Transgene | WBTransgene00000227 |
