LOCUS Exported 3906 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Encodes a HR repair template for knock-in of an EGFP insert that fused to the 5'-end of human SFPQ gene, without disturbing its promoter. Best used with px330-GFP-SFPQ vector.. ACCESSION . VERSION . KEYWORDS pUC57-GFP-SFPQ-RT SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 3906) AUTHORS Li R, Harvey AR, Hodgetts SI, Fox AH TITLE Functional dissection of NEAT1 using genome editing reveals substantial localisation of the NEAT1_1 isoform outside paraspeckles. JOURNAL RNA. 2017 Mar 21. pii: rna.059477.116. doi: 10.1261/rna.059477.116. PUBMED 28325845 REFERENCE 2 (bases 1 to 3906) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..3906 /organism="synthetic DNA construct" /mol_type="other DNA" primer_bind complement(29..51) /label=pGEX 3' /note="pGEX vectors, reverse primer" primer_bind 151..170 /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" primer_bind 364..386 /label=M13/pUC Forward /note="In lacZ gene" CDS complement(736..1452) /codon_start=1 /product="enhanced GFP" /label=EGFP /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" CDS complement(736..1452) /codon_start=1 /product="the original enhanced GFP (Yang et al., 1996)" /label=EGFP /note="mammalian codon-optimized" /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" primer_bind complement(778..799) /label=EGFP-C /note="EGFP, forward primer" primer_bind 1127..1146 /label=EXFP-R /note="For distinguishing EGFP variants, reverse primer" primer_bind 1386..1407 /label=EGFP-N /note="EGFP, reverse primer" primer_bind complement(1816..1832) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind complement(1816..1832) /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" primer_bind complement(1829..1851) /label=M13/pUC Reverse /note="In lacZ gene" protein_bind 1840..1856 /label=lac operator /bound_moiety="lac repressor encoded by lacI" /note="The lac repressor binds to the lac operator to inhibit transcription in E. coli. This inhibition can be relieved by adding lactose or isopropyl-beta-D-thiogalactopyranoside (IPTG)." promoter complement(1864..1894) /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 1909..1930 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." primer_bind complement(2047..2064) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(2218..2806) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(2298..2317) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(2984..3793) /codon_start=1 /gene="aph(3')-Ia" /product="aminoglycoside phosphotransferase" /label=KanR /note="confers resistance to kanamycin in bacteria or G418 (Geneticin(R)) in eukaryotes" /translation="MSHIQRETSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF" primer_bind 3707..3726 /label=Kan-R /note="Kanamycin resistance gene, reverse primer" primer_bind 3785..3804 /label=pENTR-R /note="pENTR vectors, reverse primer" ORIGIN 1 tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gagacggtca 61 cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg 121 ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta ctgagagtgc 181 accatatgcg gtgtgaaata ccgcacagat gcgtaaggag aaaataccgc atcaggcgcc 241 attcgccatt caggctgcgc aactgttggg aagggcgatc ggtgcgggcc tcttcgctat 301 tacgccagct ggcgaaaggg ggatgtgctg caaggcgatt aagttgggta acgccagggt 361 tttcccagtc acgacgttgt aaaacgacgg ccagagaatt cgagctcggt acctcgcgaa 421 tacatctaga tattgggtct gcggctgtgg atgcggcggc ggctgatgcg gtggcggctg 481 ctgcggcggt ggctgctgcg gtggtggctg ttgctgctgt tggtgtggag gcggtggcgg 541 gatcggaggc ttagggccgc tctggcccgg gccaggaccc atggggccgc gattctgatt 601 gaggcccatg ccgggcggcg gagaacggaa gtcgtggagg ccgccgcggc cgccgcctcc 661 tccacgcctg tggaagccac caccgccacc gccacgactc cggaaccgat cccgagacat 721 tgcagccgct gcagccttgt acagctcgtc catgccgaga gtgatcccgg cggcggtcac 781 gaactccagc aggaccatgt gatcgcgctt ctcgttgggg tctttgctca gggcggactg 841 ggtgctcagg tagtggttgt cgggcagcag cacggggccg tcgccgatgg gggtgttctg 901 ctggtagtgg tcggcgagct gcacgctgcc gtcctcgatg ttgtggcgga tcttgaagtt 961 caccttgatg ccgttcttct gcttgtcggc catgatatag acgttgtggc tgttgtagtt 1021 gtactccagc ttgtgcccca ggatgttgcc gtcctccttg aagtcgatgc ccttcagctc 1081 gatgcggttc accagggtgt cgccctcgaa cttcacctcg gcgcgggtct tgtagttgcc 1141 gtcgtccttg aagaagatgg tgcgctcctg gacgtagcct tcgggcatgg cggacttgaa 1201 gaagtcgtgc tgcttcatgt ggtcggggta gcggctgaag cactgcacgc cgtaggtcag 1261 ggtggtcacg agggtgggcc agggcacggg cagcttgccg gtggtgcaga tgaacttcag 1321 ggtcagcttg ccgtaggtgg catcgccctc gccctcgccg gacacgctga acttgtggcc 1381 gtttacgtcg ccgtccagct cgaccaggat gggcaccacc ccggtgaaca gctcctcgcc 1441 cttgctcacc atgtctgtgg tcaaggggcg gtcgaggcaa aagcgaagaa gacgctcagg 1501 aaacgtggag gccaccttgc ttctcacaaa atggcggatg acacaggcgg cgcgccgcct 1561 ttgtcctcgt cttatatagg ccagctggca cggccccgcc tactccacga tcgacagctc 1621 caatagccaa ttgcaattac tatgcttgat gacactaagt tgcgaacctc tgatcattcc 1681 tattggcccc tgggtgagag gcgggaggca tggggggact gaagtttcca gttggccagt 1741 gagcgaagaa gattgggtat cggatcccgg gcccgtcgac tgcagaggcc tgcatgcaag 1801 cttggtgtaa tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc 1861 acacaacata cgagccggaa gcataaagtg taaagcctgg ggtgcctaat gagtgagcta 1921 actcacatta attgcgttgc gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca 1981 gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc 2041 cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc 2101 tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat 2161 gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt 2221 ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg 2281 aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc 2341 tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt 2401 ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa 2461 gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta 2521 tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa 2581 caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa 2641 ctacggctac actagaagaa cagtatttgg tatctgcgct ctgctgaagc cagttacctt 2701 cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt 2761 ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat 2821 cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat 2881 gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa gttttaaatc 2941 aagcccaatc tgaataatgt tacaaccaat taaccaattc tgattagaaa aactcatcga 3001 gcatcaaatg aaactgcaat ttattcatat caggattatc aataccatat ttttgaaaaa 3061 gccgtttctg taatgaagga gaaaactcac cgaggcagtt ccataggatg gcaagatcct 3121 ggtatcggtc tgcgattccg actcgtccaa catcaataca acctattaat ttcccctcgt 3181 caaaaataag gttatcaagt gagaaatcac catgagtgac gactgaatcc ggtgagaatg 3241 gcaaaagttt atgcatttct ttccagactt gttcaacagg ccagccatta cgctcgtcat 3301 caaaatcact cgcatcaacc aaaccgttat tcattcgtga ttgcgcctga gcgagacgaa 3361 atacgcgatc gctgttaaaa ggacaattac aaacaggaat cgaatgcaac cggcgcagga 3421 acactgccag cgcatcaaca atattttcac ctgaatcagg atattcttct aatacctgga 3481 atgctgtttt tccggggatc gcagtggtga gtaaccatgc atcatcagga gtacggataa 3541 aatgcttgat ggtcggaaga ggcataaatt ccgtcagcca gtttagtctg accatctcat 3601 ctgtaacatc attggcaacg ctacctttgc catgtttcag aaacaactct ggcgcatcgg 3661 gcttcccata caagcgatag attgtcgcac ctgattgccc gacattatcg cgagcccatt 3721 tatacccata taaatcagca tccatgttgg aatttaatcg cggcctcgac gtttcccgtt 3781 gaatatggct cataacaccc cttgtattac tgtttatgta agcagacagt tttattgttc 3841 atgatgatat atttttatct tgtgcaatgt aacatcagag attttgagac acgggccaga 3901 gctgca //