LOCUS       sequence-211171-        4976 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of iRFP670-tagged Lifeact with G418 selection in 
            mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4976)
  AUTHORS   Padilla-Rodriguez M, Parker SS, Adams DG, Westerling T, Puleo JI, 
            Watson AW, Hill SM, Noon M, Gaudin R, Aaron J, Tong D, Roe DJ, 
            Knudsen B, Mouneimne G
  TITLE     The actin cytoskeletal architecture of estrogen receptor positive 
            breast cancer cells suppresses invasion.
  JOURNAL   Nat Commun. 2018 Jul 30;9(1):2980. doi: 10.1038/s41467-018-05367-2.
  PUBMED    30061623
REFERENCE   2  (bases 1 to 4976)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nat 
            Commun."; date: "2018-07-30"; volume: "9(1):2980. doi"; pages: " 
            10.1038/s41467-018-05367-2"
COMMENT     Sequence Label: Lifeact-7-iRFP670
FEATURES             Location/Qualifiers
     source          1..4976
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..304
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        305..508
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     459..479
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             575..625
                     /codon_start=1
                     /product="peptide that binds filamentous actin (Riedl et 
                     al., 2008)"
                     /label=Lifeact
                     /note="first 17 amino acids of budding yeast Abp140"
                     /translation="MGVADLIKKFESISKEE"
     regulatory      641..650
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             647..1582
                     /codon_start=1
                     /product="near-infrared fluorescent protein with an 
                     emission peak at 670 nm, engineered from a bacterial 
                     phytochrome (Shcherbakova and Verkhusha, 2013)"
                     /label=iRFP670
                     /note="derived from Rhodopseudomonas palustris RpBphP6"
                     /translation="MARKVDLTSCDREPIHIPGSIQPCGCLLACDAQAVRITRITENAG
                     AFFGRETPRVGELLADYFGETEAHALRNALAQSSDPKRPALIFGWRDGLTGRTFDISLH
                     RHDGTSIIEFEPAAAEQADNPLRLTRQIIARTKELKSLEEMAARVPRYLQAMLGYHRVM
                     LYRFADDGSGMVIGEAKRSDLESFLGQHFPASLVPQQARLLYLKNAIRVVSDSRGISSR
                     IVPEHDASGAALDLSFAHLRSISPCHLEFLRNMGVSASMSLSIIIDGTLWGLIICHHYE
                     PRAVPMAQRVAAEMFADFLSLHFTAAHHQR"
     polyA_signal    1704..1825
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1741..1760)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1795..1814
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(1832..2287)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1969..1990)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2181..2200
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2314..2418
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2420..2777
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2420..2611
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2443..2463)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2628..2763
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             2812..3606
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             2812..3606
                     /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(2866..2885)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3476..3495
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(3794..3813)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    3838..3885
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4214..4802
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4703..4722
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
       61 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      121 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      181 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      241 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      301 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      361 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      421 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      481 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcgcta
      541 ccggactcag atctcgagct caagcttcga attcatgggt gtcgcagatt tgatcaagaa
      601 attcgaaagc atctcaaagg aagaagggga tccaccggtc gccaccatgg cgcgtaaggt
      661 cgatctcacc tcctgcgatc gcgagccgat ccacatcccc ggcagcattc agccgtgcgg
      721 ctgcctgcta gcctgcgacg cgcaggcggt gcggatcacg cgcattacgg aaaatgccgg
      781 cgcgttcttt ggacgcgaaa ctccgcgggt cggtgagcta ctcgccgatt acttcggcga
      841 gaccgaagcc catgcgctgc gcaacgcact ggcgcagtcc tccgatccaa agcgaccggc
      901 gctgatcttc ggttggcgcg acggcctgac cggccgcacc ttcgacatct cactgcatcg
      961 ccatgacggt acatcgatca tcgagttcga gcctgcggcg gccgaacagg ccgacaatcc
     1021 gctgcggctg acgcggcaga tcatcgcgcg caccaaagaa ctgaagtcgc tcgaagagat
     1081 ggccgcacgg gtgccgcgct atctgcaggc gatgctcggc tatcaccgcg tgatgttgta
     1141 ccgcttcgcg gacgacggct ccgggatggt gatcggcgag gcgaagcgca gcgacctcga
     1201 gagctttctc ggtcagcact ttccggcgtc gctggtcccg cagcaggcgc ggctactgta
     1261 cttgaagaac gcgatccgcg tggtctcgga ttcgcgcggc atcagcagcc ggatcgtgcc
     1321 cgagcacgac gcctccggcg ccgcgctcga tctgtcgttc gcgcacctgc gcagcatctc
     1381 gccctgccat ctcgaatttc tgcggaacat gggcgtcagc gcctcgatgt cgctgtcgat
     1441 catcattgac ggcacgctat ggggattgat catctgtcat cattacgagc cgcgtgccgt
     1501 gccgatggcg cagcgcgtcg cggccgaaat gttcgccgac ttcttatcgc tgcacttcac
     1561 cgccgcccac caccaacgct aagcggccgc gactctagat cataatcagc cataccacat
     1621 ttgtagaggt tttacttgct ttaaaaaacc tcccacacct ccccctgaac ctgaaacata
     1681 aaatgaatgc aattgttgtt gttaacttgt ttattgcagc ttataatggt tacaaataaa
     1741 gcaatagcat cacaaatttc acaaataaag catttttttc actgcattct agttgtggtt
     1801 tgtccaaact catcaatgta tcttaaggcg taaattgtaa gcgttaatat tttgttaaaa
     1861 ttcgcgttaa atttttgtta aatcagctca ttttttaacc aataggccga aatcggcaaa
     1921 atcccttata aatcaaaaga atagaccgag atagggttga gtgttgttcc agtttggaac
     1981 aagagtccac tattaaagaa cgtggactcc aacgtcaaag ggcgaaaaac cgtctatcag
     2041 ggcgatggcc cactacgtga accatcaccc taatcaagtt ttttggggtc gaggtgccgt
     2101 aaagcactaa atcggaaccc taaagggagc ccccgattta gagcttgacg gggaaagccg
     2161 gcgaacgtgg cgagaaagga agggaagaaa gcgaaaggag cgggcgctag ggcgctggca
     2221 agtgtagcgg tcacgctgcg cgtaaccacc acacccgccg cgcttaatgc gccgctacag
     2281 ggcgcgtcag gtggcacttt tcggggaaat gtgcgcggaa cccctatttg tttatttttc
     2341 taaatacatt caaatatgta tccgctcatg agacaataac cctgataaat gcttcaataa
     2401 tattgaaaaa ggaagagtcc tgaggcggaa agaaccagct gtggaatgtg tgtcagttag
     2461 ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt
     2521 agtcagcaac caggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca
     2581 tgcatctcaa ttagtcagca accatagtcc cgcccctaac tccgcccatc ccgcccctaa
     2641 ctccgcccag ttccgcccat tctccgcccc atggctgact aatttttttt atttatgcag
     2701 aggccgaggc cgcctcggcc tctgagctat tccagaagta gtgaggaggc ttttttggag
     2761 gcctaggctt ttgcaaagat cgatcaagag acaggatgag gatcgtttcg catgattgaa
     2821 caagatggat tgcacgcagg ttctccggcc gcttgggtgg agaggctatt cggctatgac
     2881 tgggcacaac agacaatcgg ctgctctgat gccgccgtgt tccggctgtc agcgcagggg
     2941 cgcccggttc tttttgtcaa gaccgacctg tccggtgccc tgaatgaact gcaagacgag
     3001 gcagcgcggc tatcgtggct ggccacgacg ggcgttcctt gcgcagctgt gctcgacgtt
     3061 gtcactgaag cgggaaggga ctggctgcta ttgggcgaag tgccggggca ggatctcctg
     3121 tcatctcacc ttgctcctgc cgagaaagta tccatcatgg ctgatgcaat gcggcggctg
     3181 catacgcttg atccggctac ctgcccattc gaccaccaag cgaaacatcg catcgagcga
     3241 gcacgtactc ggatggaagc cggtcttgtc gatcaggatg atctggacga agagcatcag
     3301 gggctcgcgc cagccgaact gttcgccagg ctcaaggcga gcatgcccga cggcgaggat
     3361 ctcgtcgtga cccatggcga tgcctgcttg ccgaatatca tggtggaaaa tggccgcttt
     3421 tctggattca tcgactgtgg ccggctgggt gtggcggacc gctatcagga catagcgttg
     3481 gctacccgtg atattgctga agagcttggc ggcgaatggg ctgaccgctt cctcgtgctt
     3541 tacggtatcg ccgctcccga ttcgcagcgc atcgccttct atcgccttct tgacgagttc
     3601 ttctgagcgg gactctgggg ttcgaaatga ccgaccaagc gacgcccaac ctgccatcac
     3661 gagatttcga ttccaccgcc gccttctatg aaaggttggg cttcggaatc gttttccggg
     3721 acgccggctg gatgatcctc cagcgcgggg atctcatgct ggagttcttc gcccacccta
     3781 gggggaggct aactgaaaca cggaaggaga caataccgga aggaacccgc gctatgacgg
     3841 caataaaaag acagaataaa acgcacggtg ttgggtcgtt tgttcataaa cgcggggttc
     3901 ggtcccaggg ctggcactct gtcgataccc caccgagacc ccattggggc caatacgccc
     3961 gcgtttcttc cttttcccca ccccaccccc caagttcggg tgaaggccca gggctcgcag
     4021 ccaacgtcgg ggcggcaggc cctgccatag cctcaggtta ctcatatata ctttagattg
     4081 atttaaaact tcatttttaa tttaaaagga tctaggtgaa gatccttttt gataatctca
     4141 tgaccaaaat cccttaacgt gagttttcgt tccactgagc gtcagacccc gtagaaaaga
     4201 tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa
     4261 aaccaccgct accagcggtg gtttgtttgc cggatcaaga gctaccaact ctttttccga
     4321 aggtaactgg cttcagcaga gcgcagatac caaatactgt tcttctagtg tagccgtagt
     4381 taggccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg ctaatcctgt
     4441 taccagtggc tgctgccagt ggcgataagt cgtgtcttac cgggttggac tcaagacgat
     4501 agttaccgga taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca cagcccagct
     4561 tggagcgaac gacctacacc gaactgagat acctacagcg tgagctatga gaaagcgcca
     4621 cgcttcccga agggagaaag gcggacaggt atccggtaag cggcagggtc ggaacaggag
     4681 agcgcacgag ggagcttcca gggggaaacg cctggtatct ttatagtcct gtcgggtttc
     4741 gccacctctg acttgagcgt cgatttttgt gatgctcgtc aggggggcgg agcctatgga
     4801 aaaacgccag caacgcggcc tttttacggt tcctggcctt ttgctggcct tttgctcaca
     4861 tgttctttcc tgcgttatcc cctgattctg tggataaccg tattaccgcc atgcattagt
     4921 tattaatagt aatcaattac ggggtcatta gttcatagcc catatatgga gttccg
//
