LOCUS Exported 5219 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Encodes a HR repair template that mutates the poly(A) signal as well as a few splicing factor binding sites of human NEAT1_1 isoform. Best used with px335-NEAT1_IS_v1 and v2 vectors.. ACCESSION . VERSION . KEYWORDS pYFP-NEAT1_IS_RT SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5219) 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 5219) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..5219 /organism="synthetic DNA construct" /mol_type="other DNA" primer_bind 180..199 /label=EXFP-R /note="For distinguishing EGFP variants, reverse primer" primer_bind 439..460 /label=EGFP-N /note="EGFP, reverse primer" regulatory 502..511 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" promoter complement(550..753) /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind complement(579..599) /label=CMV-F /note="Human CMV immediate early promoter, forward primer" enhancer 754..1057 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" rep_origin complement(1232..1820) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(1312..1331) /label=pBR322ori-F /note="pBR322 origin, forward primer" polyA_signal 2149..2196 /label=HSV TK poly(A) signal /note="herpes simplex virus thymidine kinase polyadenylation signal (Cole and Stacy, 1985)" primer_bind 2221..2240 /label=TK-pA-R /note="Thymidine kinase polyA, reverse primer" CDS complement(2428..3222) /codon_start=1 /gene="aph(3')-II (or nptII)" /product="aminoglycoside phosphotransferase from Tn5" /label=NeoR/KanR /note="confers resistance to neomycin, kanamycin, and G418 (Geneticin(R))" /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF" primer_bind complement(2539..2558) /label=Neo-F /note="Neomycin resistance gene, forward primer" primer_bind 3149..3168 /label=Neo-R /note="Neomycin resistance gene, reverse primer" promoter complement(3257..3614) /label=SV40 promoter /note="SV40 enhancer and early promoter" rep_origin 3271..3406 /label=SV40 ori /note="SV40 origin of replication" primer_bind complement(3325..3344) /label=SV40pro-F /note="SV40 promoter/origin, forward primer" primer_bind 3571..3591 /label=pBABE 3' /note="SV40 enhancer, reverse primer for pBABE vectors" promoter complement(3616..3720) /gene="bla" /label=AmpR promoter rep_origin 3747..4202 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(3834..3853) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 4044..4065 /label=F1ori-F /note="F1 origin, forward primer" polyA_signal 4697..4818 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(4708..4727) /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind 4762..4781 /label=SV40pA-R /note="SV40 polyA, reverse primer" misc_feature 4942..5007 /label=MCS /note="multiple cloning site" CDS complement(5008..505) /codon_start=1 /product="enhanced YFP" /label=EYFP /note="mammalian codon-optimized" /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" primer_bind complement(5050..5071) /label=EGFP-C /note="EGFP, forward primer" ORIGIN 1 ggatcttgaa gttcaccttg atgccgttct tctgcttgtc ggccatgata tagacgttgt 61 ggctgttgta gttgtactcc agcttgtgcc ccaggatgtt gccgtcctcc ttgaagtcga 121 tgcccttcag ctcgatgcgg ttcaccaggg tgtcgccctc gaacttcacc tcggcgcggg 181 tcttgtagtt gccgtcgtcc ttgaagaaga tggtgcgctc ctggacgtag ccttcgggca 241 tggcggactt gaagaagtcg tgctgcttca tgtggtcggg gtagcgggcg aagcactgca 301 ggccgtagcc gaaggtggtc acgagggtgg gccagggcac gggcagcttg ccggtggtgc 361 agatgaactt cagggtcagc ttgccgtagg tggcatcgcc ctcgccctcg ccggacacgc 421 tgaacttgtg gccgtttacg tcgccgtcca gctcgaccag gatgggcacc accccggtga 481 acagctcctc gcccttgctc accatggtgg cgaccggtag cgctagcgga tctgacggtt 541 cactaaacca gctctgctta tatagacctc ccaccgtaca cgcctaccgc ccatttgcgt 601 caatggggcg gagttgttac gacattttgg aaagtcccgt tgattttggt gccaaaacaa 661 actcccattg acgtcaatgg ggtggagact tggaaatccc cgtgagtcaa accgctatcc 721 acgcccattg atgtactgcc aaaaccgcat caccatggta atagcgatga ctaatacgta 781 gatgtactgc caagtaggaa agtcccataa ggtcatgtac tgggcataat gccaggcggg 841 ccatttaccg tcattgacgt caataggggg cgtacttggc atatgataca cttgatgtac 901 tgccaagtgg gcagtttacc gtaaatactc cacccattga cgtcaatgga aagtccctat 961 tggcgttact atgggaacat acgtcattat tgacgtcaat gggcgggggt cgttgggcgg 1021 tcagccaggc gggccattta ccgtaagtta tgtaacgcgg aactccatat atgggctatg 1081 aactaatgac cccgtaattg attactatta ataactaatg catggcggta atacggttat 1141 ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag caaaaggcca 1201 ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc cctgacgagc 1261 atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc 1321 aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg 1381 gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta 1441 ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg 1501 ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac 1561 acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag 1621 gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga agaacagtat 1681 ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat 1741 ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc 1801 gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt 1861 ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg atcttcacct 1921 agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat gagtaacctg 1981 aggctatggc agggcctgcc gccccgacgt tggctgcgag ccctgggcct tcacccgaac 2041 ttggggggtg gggtggggaa aaggaagaaa cgcgggcgta ttggccccaa tggggtctcg 2101 gtggggtatc gacagagtgc cagccctggg accgaacccc gcgtttatga acaaacgacc 2161 caacaccgtg cgttttattc tgtcttttta ttgccgtcat agcgcgggtt ccttccggta 2221 ttgtctcctt ccgtgtttca gttagcctcc ccctagggtg ggcgaagaac tccagcatga 2281 gatccccgcg ctggaggatc atccagccgg cgtcccggaa aacgattccg aagcccaacc 2341 tttcatagaa ggcggcggtg gaatcgaaat ctcgtgatgg caggttgggc gtcgcttggt 2401 cggtcatttc gaaccccaga gtcccgctca gaagaactcg tcaagaaggc gatagaaggc 2461 gatgcgctgc gaatcgggag cggcgatacc gtaaagcacg aggaagcggt cagcccattc 2521 gccgccaagc tcttcagcaa tatcacgggt agccaacgct atgtcctgat agcggtccgc 2581 cacacccagc cggccacagt cgatgaatcc agaaaagcgg ccattttcca ccatgatatt 2641 cggcaagcag gcatcgccat gggtcacgac gagatcctcg ccgtcgggca tgctcgcctt 2701 gagcctggcg aacagttcgg ctggcgcgag cccctgatgc tcttcgtcca gatcatcctg 2761 atcgacaaga ccggcttcca tccgagtacg tgctcgctcg atgcgatgtt tcgcttggtg 2821 gtcgaatggg caggtagccg gatcaagcgt atgcagccgc cgcattgcat cagccatgat 2881 ggatactttc tcggcaggag caaggtgaga tgacaggaga tcctgccccg gcacttcgcc 2941 caatagcagc cagtcccttc ccgcttcagt gacaacgtcg agcacagctg cgcaaggaac 3001 gcccgtcgtg gccagccacg atagccgcgc tgcctcgtct tgcagttcat tcagggcacc 3061 ggacaggtcg gtcttgacaa aaagaaccgg gcgcccctgc gctgacagcc ggaacacggc 3121 ggcatcagag cagccgattg tctgttgtgc ccagtcatag ccgaatagcc tctccaccca 3181 agcggccgga gaacctgcgt gcaatccatc ttgttcaatc atgcgaaacg atcctcatcc 3241 tgtctcttga tcgatctttg caaaagccta ggcctccaaa aaagcctcct cactacttct 3301 ggaatagctc agaggccgag gcggcctcgg cctctgcata aataaaaaaa attagtcagc 3361 catggggcgg agaatgggcg gaactgggcg gagttagggg cgggatgggc ggagttaggg 3421 gcgggactat ggttgctgac taattgagat gcatgctttg catacttctg cctgctgggg 3481 agcctgggga ctttccacac ctggttgctg actaattgag atgcatgctt tgcatacttc 3541 tgcctgctgg ggagcctggg gactttccac accctaactg acacacattc cacagctggt 3601 tctttccgcc tcaggactct tcctttttca atattattga agcatttatc agggttattg 3661 tctcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg 3721 cacatttccc cgaaaagtgc cacctgacgc gccctgtagc ggcgcattaa gcgcggcggg 3781 tgtggtggtt acgcgcagcg tgaccgctac acttgccagc gccctagcgc ccgctccttt 3841 cgctttcttc ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg 3901 ggggctccct ttagggttcc gatttagtgc tttacggcac ctcgacccca aaaaacttga 3961 ttagggtgat ggttcacgta gtgggccatc gccctgatag acggtttttc gccctttgac 4021 gttggagtcc acgttcttta atagtggact cttgttccaa actggaacaa cactcaaccc 4081 tatctcggtc tattcttttg atttataagg gattttgccg atttcggcct attggttaaa 4141 aaatgagctg atttaacaaa aatttaacgc gaattttaac aaaatattaa cgcttacaat 4201 ttacgccggt taccatgctc tcctacatgg ccttaattac ttggagctag caaatctaga 4261 cctaaatcta caatatgaag atacatccaa agtcgttatg aaggcaatgt gataggggtc 4321 gagaaatgta acatagcaat acaacaattc taatgagttt agaactcaaa ccttgtctgt 4381 gctgtaaagc aaggggaaga aaagtgatta gtaattatgt cgatgacgta acagaattag 4441 ttcttaccat cgagagcaac ataccagtac tttcaaccat ctacttttcg agtttcaatt 4501 tcacaacagc atacccgaga ctacttaacc atacatgcgt gactaataca ctctaaatcc 4561 caacgacagt aattgtttgc atcatcccca agtcattggt taagaactga actaccatgt 4621 caacatattt ttaaacattt aaaaaatgga acccagaaga cagaaagatc ccaacgataa 4681 aaataaccat gattgttaag atacattgat gagtttggac aaaccacaac tagaatgcag 4741 tgaaaaaaat gctttatttg tgaaatttgt gatgctattg ctttatttgt aaccattata 4801 agctgcaata aacaagttaa caacaacaat tgcattcatt ttatgtttca ggttcagggg 4861 gaggtgtggg aggtttttta aagcaagtaa aacctctaca aatgtggtat ggctgattat 4921 gatcagttat ctagatccgg tggatcccgg gcccgcggta ccgtcgactg cagaattcga 4981 agcttgagct cgagatctga gtccggactt gtacagctcg tccatgccga gagtgatccc 5041 ggcggcggtc acgaactcca gcaggaccat gtgatcgcgc ttctcgttgg ggtctttgct 5101 cagggcggac tggtagctca ggtagtggtt gtcgggcagc agcacggggc cgtcgccgat 5161 gggggtgttc tgctggtagt ggtcggcgag ctgcacgctg ccgtcctcga tgttgtggc //