LOCUS Exported 5932 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Encodes a HR repair template for knock-in a YFP expression cassette at the promoter of human NEAT1 gene. Best used with px330-NEAT1pr_v1 vector, but also works with px330-NEAT1pr_v2.. ACCESSION . VERSION . KEYWORDS pYFP-NEAT1pr_v1-RT SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5932) TITLE Unpublished Fox lab plasmids REFERENCE 2 (bases 1 to 5932) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..5932 /organism="synthetic DNA construct" /mol_type="other DNA" enhancer 703..1006 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 1007..1210 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 1161..1181 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" regulatory 1249..1258 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 1255..1971 /codon_start=1 /product="enhanced YFP" /label=EYFP /note="mammalian codon-optimized" /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" primer_bind complement(1300..1321) /label=EGFP-N /note="EGFP, reverse primer" primer_bind complement(1561..1580) /label=EXFP-R /note="For distinguishing EGFP variants, reverse primer" primer_bind 1908..1929 /label=EGFP-C /note="EGFP, forward primer" misc_feature 1972..2037 /label=MCS /note="multiple cloning site" polyA_signal 2161..2282 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(2198..2217) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 2252..2271 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" rep_origin complement(2848..3303) /direction=LEFT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(2985..3006) /label=F1ori-F /note="F1 origin, forward primer" primer_bind 3197..3216 /label=F1ori-R /note="F1 origin, reverse primer" promoter 3330..3434 /gene="bla" /label=AmpR promoter promoter 3436..3793 /label=SV40 promoter /note="SV40 enhancer and early promoter" primer_bind complement(3459..3479) /label=pBABE 3' /note="SV40 enhancer, reverse primer for pBABE vectors" rep_origin 3644..3779 /label=SV40 ori /note="SV40 origin of replication" primer_bind 3706..3725 /label=SV40pro-F /note="SV40 promoter/origin, forward primer" CDS 3828..4622 /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(3882..3901) /label=Neo-R /note="Neomycin resistance gene, reverse primer" primer_bind 4492..4511 /label=Neo-F /note="Neomycin resistance gene, forward primer" primer_bind complement(4810..4829) /label=TK-pA-R /note="Thymidine kinase polyA, reverse primer" polyA_signal 4854..4901 /label=HSV TK poly(A) signal /note="herpes simplex virus thymidine kinase polyadenylation signal (Cole and Stacy, 1985)" rep_origin 5230..5818 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 5719..5738 /label=pBR322ori-F /note="pBR322 origin, forward primer" ORIGIN 1 tagttatgga cgctatcagc ccgcagcagg gtttcctggc cagagaaacc gcctgttggg 61 gtgcggagcg ttccacgtcc cctcgcagca ccccgctggc cttctgggcc gccccgcccc 121 caacccgcgc ccgggcgctg acgtcatcgg ccgagcccga ctcggaacca ccgcccgaaa 181 gtcacgcgcg cctcccggcg ggacgtgccc tctgcagaga tccctccgcc gccgcctgga 241 attttccaga tgtcctgccg gcccttcttt cgggcctcgg ctgggcgtgg tgacctcaag 301 cgaccccggc ggctcccgtc gcccactcaa gagggcgccc ccgcccgcca cgcccccagc 361 ccctctcggg gaagcgcgtc ccccgcccga cctcaacaac atccgggaag aaaaggggtc 421 ttcttcctca tggcattcgc ctcccaaatg tcaccttgag cgctgtccgc gatcccaaaa 481 agcactgtta aagagaagcg gggatacact ggggtccttg cgtgggggcc gcctgggaga 541 ccatgcaccg cccgggagtc tctccgggca gggtcgggga gggacttttt tgccggtggc 601 cgtggaggaa tcgtcccgtt gagcaatgac cccggtgacg cggctgaggg catagtaatc 661 aattacgggg tcattagttc atagcccata tatggagttc cgcgttacat aacttacggt 721 aaatggcccg cctggctgac cgcccaacga cccccgccca ttgacgtcaa taatgacgta 781 tgttcccata gtaacgccaa tagggacttt ccattgacgt caatgggtgg agtatttacg 841 gtaaactgcc cacttggcag tacatcaagt gtatcatatg ccaagtacgc cccctattga 901 cgtcaatgac ggtaaatggc ccgcctggca ttatgcccag tacatgacct tatgggactt 961 tcctacttgg cagtacatct acgtattagt catcgctatt accatggtga tgcggttttg 1021 gcagtacatc aatgggcgtg gatagcggtt tgactcacgg ggatttccaa gtctccaccc 1081 cattgacgtc aatgggagtt tgttttggca ccaaaatcaa cgggactttc caaaatgtcg 1141 taacaactcc gccccattga cgcaaatggg cggtaggcgt gtacggtggg aggtctatat 1201 aagcagagct ggtttagtga accgtcagat ccgctagcgc taccggtcgc caccatggtg 1261 agcaagggcg aggagctgtt caccggggtg gtgcccatcc tggtcgagct ggacggcgac 1321 gtaaacggcc acaagttcag cgtgtccggc gagggcgagg gcgatgccac ctacggcaag 1381 ctgaccctga agttcatctg caccaccggc aagctgcccg tgccctggcc caccctcgtg 1441 accaccttcg gctacggcct gcagtgcttc gcccgctacc ccgaccacat gaagcagcac 1501 gacttcttca agtccgccat gcccgaaggc tacgtccagg agcgcaccat cttcttcaag 1561 gacgacggca actacaagac ccgcgccgag gtgaagttcg agggcgacac cctggtgaac 1621 cgcatcgagc tgaagggcat cgacttcaag gaggacggca acatcctggg gcacaagctg 1681 gagtacaact acaacagcca caacgtctat atcatggccg acaagcagaa gaacggcatc 1741 aaggtgaact tcaagatccg ccacaacatc gaggacggca gcgtgcagct cgccgaccac 1801 taccagcaga acacccccat cggcgacggc cccgtgctgc tgcccgacaa ccactacctg 1861 agctaccagt ccgccctgag caaagacccc aacgagaagc gcgatcacat ggtcctgctg 1921 gagttcgtga ccgccgccgg gatcactctc ggcatggacg agctgtacaa gtccggactc 1981 agatctcgag ctcaagcttc gaattctgca gtcgacggta ccgcgggccc gggatccacc 2041 ggatctagat aactgatcat aatcagccat accacatttg tagaggtttt acttgcttta 2101 aaaaacctcc cacacctccc cctgaacctg aaacataaaa tgaatgcaat tgttgttgtt 2161 aacttgttta ttgcagctta taatggttac aaataaagca atagcatcac aaatttcaca 2221 aataaagcat ttttttcact gcattctagt tgtggtttgt ccaaactcat caatgtatct 2281 taattgtggc aagtccagcc ggagttagcg acagggaggg atgcgcgcct gggtgtagtt 2341 gtgggggagg aagtggctag ctcagggctt caggggacag acagggagag atgactgagt 2401 tagatgagac gagggggcgg gctgggggtg cgagaaggaa gcttggcaag gagactaggt 2461 ctagggggac cacagtgggg caggctgcat ggaaaatatc cgcagggtcc cccaggcaga 2521 acagccacgc tccaggccag gctgtcccta ctgcctggtg gagggggaac ttgacctctg 2581 ggagggcgcc gctcttgcat agctgagcga gcccgggtgc gctggtctgt gtggaaggag 2641 gaaggcaggg agaggtagaa ggggtggagg agtcaggagg aataggccgc agcagccctg 2701 gaaatgatca ggaaggcagg cagtgggtgc agggctgcag gagggccggg agggctaatc 2761 ttcaacttgt ccatgccagc agcccctttt tttccagacc aagggctgtg aacccgcctg 2821 gggatgaggc ctggtcttgt ggaacttaaa ttgtaagcgt taatattttg ttaaaattcg 2881 cgttaaattt ttgttaaatc agctcatttt ttaaccaata ggccgaaatc ggcaaaatcc 2941 cttataaatc aaaagaatag accgagatag ggttgagtgt tgttccagtt tggaacaaga 3001 gtccactatt aaagaacgtg gactccaacg tcaaagggcg aaaaaccgtc tatcagggcg 3061 atggcccact acgtgaacca tcaccctaat caagtttttt ggggtcgagg tgccgtaaag 3121 cactaaatcg gaaccctaaa gggagccccc gatttagagc ttgacgggga aagccggcga 3181 acgtggcgag aaaggaaggg aagaaagcga aaggagcggg cgctagggcg ctggcaagtg 3241 tagcggtcac gctgcgcgta accaccacac ccgccgcgct taatgcgccg ctacagggcg 3301 cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc tatttgttta tttttctaaa 3361 tacattcaaa tatgtatccg ctcatgagac aataaccctg ataaatgctt caataatatt 3421 gaaaaaggaa gagtcctgag gcggaaagaa ccagctgtgg aatgtgtgtc agttagggtg 3481 tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc 3541 agcaaccagg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc aaagcatgca 3601 tctcaattag tcagcaacca tagtcccgcc cctaactccg cccatcccgc ccctaactcc 3661 gcccagttcc gcccattctc cgccccatgg ctgactaatt ttttttattt atgcagaggc 3721 cgaggccgcc tcggcctctg agctattcca gaagtagtga ggaggctttt ttggaggcct 3781 aggcttttgc aaagatcgat caagagacag gatgaggatc gtttcgcatg attgaacaag 3841 atggattgca cgcaggttct ccggccgctt gggtggagag gctattcggc tatgactggg 3901 cacaacagac aatcggctgc tctgatgccg ccgtgttccg gctgtcagcg caggggcgcc 3961 cggttctttt tgtcaagacc gacctgtccg gtgccctgaa tgaactgcaa gacgaggcag 4021 cgcggctatc gtggctggcc acgacgggcg ttccttgcgc agctgtgctc gacgttgtca 4081 ctgaagcggg aagggactgg ctgctattgg gcgaagtgcc ggggcaggat ctcctgtcat 4141 ctcaccttgc tcctgccgag aaagtatcca tcatggctga tgcaatgcgg cggctgcata 4201 cgcttgatcc ggctacctgc ccattcgacc accaagcgaa acatcgcatc gagcgagcac 4261 gtactcggat ggaagccggt cttgtcgatc aggatgatct ggacgaagag catcaggggc 4321 tcgcgccagc cgaactgttc gccaggctca aggcgagcat gcccgacggc gaggatctcg 4381 tcgtgaccca tggcgatgcc tgcttgccga atatcatggt ggaaaatggc cgcttttctg 4441 gattcatcga ctgtggccgg ctgggtgtgg cggaccgcta tcaggacata gcgttggcta 4501 cccgtgatat tgctgaagag cttggcggcg aatgggctga ccgcttcctc gtgctttacg 4561 gtatcgccgc tcccgattcg cagcgcatcg ccttctatcg ccttcttgac gagttcttct 4621 gagcgggact ctggggttcg aaatgaccga ccaagcgacg cccaacctgc catcacgaga 4681 tttcgattcc accgccgcct tctatgaaag gttgggcttc ggaatcgttt tccgggacgc 4741 cggctggatg atcctccagc gcggggatct catgctggag ttcttcgccc accctagggg 4801 gaggctaact gaaacacgga aggagacaat accggaagga acccgcgcta tgacggcaat 4861 aaaaagacag aataaaacgc acggtgttgg gtcgtttgtt cataaacgcg gggttcggtc 4921 ccagggctgg cactctgtcg ataccccacc gagaccccat tggggccaat acgcccgcgt 4981 ttcttccttt tccccacccc accccccaag ttcgggtgaa ggcccagggc tcgcagccaa 5041 cgtcggggcg gcaggccctg ccatagcctc aggttactca tatatacttt agattgattt 5101 aaaacttcat ttttaattta aaaggatcta ggtgaagatc ctttttgata atctcatgac 5161 caaaatccct taacgtgagt tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa 5221 aggatcttct tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc 5281 accgctacca gcggtggttt gtttgccgga tcaagagcta ccaactcttt ttccgaaggt 5341 aactggcttc agcagagcgc agataccaaa tactgtcctt ctagtgtagc cgtagttagg 5401 ccaccacttc aagaactctg tagcaccgcc tacatacctc gctctgctaa tcctgttacc 5461 agtggctgct gccagtggcg ataagtcgtg tcttaccggg ttggactcaa gacgatagtt 5521 accggataag gcgcagcggt cgggctgaac ggggggttcg tgcacacagc ccagcttgga 5581 gcgaacgacc tacaccgaac tgagatacct acagcgtgag ctatgagaaa gcgccacgct 5641 tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa caggagagcg 5701 cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca 5761 cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa 5821 cgccagcaac gcggcctttt tacggttcct ggccttttgc tggccttttg ctcacatgtt 5881 ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgccatgc at //