LOCUS       sequence-474530-        8353 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses human IARS2 with mito-location signal replaced by flag tag
            in mammalian cells, also carrying synonymous mutations described in 
            Addgene #232340.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8353)
  TITLE     Mechanism of Isoleucyl-tRNA synthetase 2 regulating proliferation 
            and apoptosis of cervical cancer cells
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 8353)
  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: 
            "Unpublished"
COMMENT     Sequence Label: pcDNA3.1-flag-IARS2-Delta-1-48
FEATURES             Location/Qualifiers
     source          1..8353
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             934..957
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     RBS             1179..1187
                     /label=Shine-Dalgarno sequence
                     /note="full consensus sequence for ribosome-binding sites 
                     upstream of start codons in E. coli; complementary to a 
                     region in the 3' end of the 16S rRNA (Chen et al., 1994)"
     RBS             2200..2211
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     primer_bind     complement(3944..3961)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3950..4174
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      4220..4648
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4307..4326)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4517..4538
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4657..4677)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4662..4991
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4842..4977
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4904..4923
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             5058..5852
                     /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
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(5112..5131)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5722..5741
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    6026..6147
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6063..6082)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6117..6136
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(6196..6212)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6196..6212)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(6209..6231)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    6220..6236
                     /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(join(6244..6250,6251..6268,6269..6274))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6289..6310
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6427..6444)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6598..7186)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6678..6697)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7357..8148,8149..8217))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     7980..7999
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(8218..8322)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gctggctagc
      901 gtttaaacgg gccctctaga ctcgaccacc atggactaca aggacgacga tgataagggc
      961 ggatccctcg accggaccat gagtaaccac cagccgaact cgaatagtgg cagataccgg
     1021 gacacggtgc tgctgccgca gacgagcttc cccatgaagc tgctgggccg ccagcagccg
     1081 gacacggagc tggagatcca gcagaaatgt ggattttcag aactttattc atggcaaaga
     1141 gaaagaaaag taaagacaga attttgcctt catgatggac ctccttatgc aaacggtgac
     1201 cctcatgttg gacatgcttt aaataagatt ttgaaagaca tagccaatcg attccatatg
     1261 atgaatggct ccaaaataca ttttgtgccc ggctgggatt gtcatgggtt gcccattgaa
     1321 ataaaagtat tatcagaact tggtagagaa gctcagaatc tttcagctat ggaaattaga
     1381 aagaaagcta gatcatttgc taaagcagcc attgagaaac agaaatcagc atttattcgt
     1441 tggggaataa tggcagattg gaataattgc tactatacat ttgatgggaa gtatgaagcc
     1501 aaacagttga gaacttttta ccaaatgtat gataagggct tggtttatcg atcttacaaa
     1561 cctgtgtttt ggtctccgtc atctaggact gcattggctg aagcagaact tgaatataat
     1621 cctgagcatg tcagtcgttc aatatatgta aaatttcctc tcttaaagcc ttctccaaaa
     1681 ttggcatctc ttatagatgg ttcatctcct gttagtattt tggtctggac cacacaacct
     1741 tggacgattc cagccaatga agctgtttgc tatatgcctg aatcaaagta tgctgttgtg
     1801 aaatgttcta agtctggaga cctctacgta ctggcggcag ataaagtagc atctgttgct
     1861 tctactttgg aaacaacatt tgagactatt tcaacacttt caggtgtaga tttggaaaat
     1921 ggtacttgca gccaccctct gattcctgat aaagcctctc ctcttttacc tgcaaatcat
     1981 gtgaccatgg caaaaggaac gggattggtt cacacagccc cagctcatgg tatggaagac
     2041 tacggtgtag cgtctcagca caacctgccc atggattgtc tagtggacga agatggagtt
     2101 ttcacagatg ttgcaggtcc tgaacttcaa aacaaggctg tccttgaaga gggaactgat
     2161 gtggttataa agatgcttca gactgcaaag aatttgttga aagaggagaa attggtgcat
     2221 agctatccgt atgactggag gaccaagaaa cctgtggtta ttcgtgccag caagcagtgg
     2281 tttataaaca tcacggatat taagactgca gccaaggaat tgttaaaaaa ggtgaaattt
     2341 attcctggat cagcactgaa tggcatggtt gaaatgatgg acaggcggcc atattggtgt
     2401 atatcaaggc aaagagtttg gggtgttcca attcctgtgt ttcatcataa gaccaaggat
     2461 gaatacttga tcaacagcca aaccactgag catattgtta aactagtgga acaacacggc
     2521 agtgatatct ggtggactct tccccctgaa caacttcttc caaaagaagt cttatctgag
     2581 gttggtggcc ctgatgcctt ggaatatgtg ccaggtcagg atattttgga catctggttt
     2641 gatagcggaa cttcatggtc ttatgttctt ccaggtcctg accaaagagc agatttgtac
     2701 ttggaaggaa aagaccagct cgggggttgg tttcagtcat ccttattaac aagtgtggca
     2761 gcaaggaaga gagcacctta taagacagtg attgttcatg gatttaccct tggagaaaag
     2821 ggagaaaaga tgtccaagtc tcttgggaat gtcattcatc ctgatgttgt cgttaatgga
     2881 ggacaagatc aaagcaaaga gcctccgtat ggtgctgatg tccttcgctg gtgggtagct
     2941 gattccaatg tcttcaccga agttgcaatt ggcccatccg tgctcaatgc tgccagagat
     3001 gatattagca agcttaggaa tacacttcgc tttcttttgg gaaatgtggc tgatttcaac
     3061 ccagaaacag attccatccc tgtaaacgat atgtatgtca tagaccagta catgctacac
     3121 ttactgcagg atttggcaaa caagattacc gaattataca aacaatatga ttttggaaaa
     3181 gttgttcggc tgttacggac gttttatacc agagagctct ctaactttta tttcagtata
     3241 atcaaagata ggctctattg tgaaaaggaa aatgacccca aacgacgctc ttgtcagact
     3301 gcattagttg aaattttgga tgtaatagtt cgttcttttg ctcccattct tcctcacctg
     3361 gctgaagagg tgttccagca cataccttat attaaagagc ccaagagtgt tttccgtact
     3421 gggtggatta gtactagttc tatctggaaa aagcccgggt tggaagaagc tgtggagagt
     3481 gcgtgtgcaa tgcgagactc atttcttgga agcatccctg gcaaaaatgc agctgagtac
     3541 aaggttatca ctgtgataga acctggactg ctttttgaga taatagagat gctgcagtct
     3601 gaagagactt ccagcacctc tcagttgaat gaattaatga tggcttctga gtcaacttta
     3661 ctggctcagg aaccacgaga gatgactgca gatgtaatcg agcttaaagg gaaattcctc
     3721 atcaacttag aaggtggtga tattcgtgaa gagtcttcct ataaagtaat tgtcatgccg
     3781 actacgaaag aaaaatgccc ccgttgttgg aagtatacag cggagtcttc agatacactg
     3841 tgtcctcgat gtgcagaagt tgtcagtgga aaataggaat tccaccacac tggactagtg
     3901 gatccgagct cggtaccaag cttaagttta aaccgctgat cagcctcgac tgtgccttct
     3961 agttgccagc catctgttgt ttgcccctcc cccgtgcctt ccttgaccct ggaaggtgcc
     4021 actcccactg tcctttccta ataaaatgag gaaattgcat cgcattgtct gagtaggtgt
     4081 cattctattc tggggggtgg ggtggggcag gacagcaagg gggaggattg ggaagacaat
     4141 agcaggcatg ctggggatgc ggtgggctct atggcttctg aggcggaaag aaccagctgg
     4201 ggctctaggg ggtatcccca cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg
     4261 gttacgcgca gcgtgaccgc tacacttgcc agcgccctag cgcccgctcc tttcgctttc
     4321 ttcccttcct ttctcgccac gttcgccggc tttccccgtc aagctctaaa tcgggggctc
     4381 cctttagggt tccgatttag tgctttacgg cacctcgacc ccaaaaaact tgattagggt
     4441 gatggttcac gtagtgggcc atcgccctga tagacggttt ttcgcccttt gacgttggag
     4501 tccacgttct ttaatagtgg actcttgttc caaactggaa caacactcaa ccctatctcg
     4561 gtctattctt ttgatttata agggattttg ccgatttcgg cctattggtt aaaaaatgag
     4621 ctgatttaac aaaaatttaa cgcgaattaa ttctgtggaa tgtgtgtcag ttagggtgtg
     4681 gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag
     4741 caaccaggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc
     4801 tcaattagtc agcaaccata gtcccgcccc taactccgcc catcccgccc ctaactccgc
     4861 ccagttccgc ccattctccg ccccatggct gactaatttt ttttatttat gcagaggccg
     4921 aggccgcctc tgcctctgag ctattccaga agtagtgagg aggctttttt ggaggcctag
     4981 gcttttgcaa aaagctcccg ggagcttgta tatccatttt cggatctgat caagagacag
     5041 gatgaggatc gtttcgcatg attgaacaag atggattgca cgcaggttct ccggccgctt
     5101 gggtggagag gctattcggc tatgactggg cacaacagac aatcggctgc tctgatgccg
     5161 ccgtgttccg gctgtcagcg caggggcgcc cggttctttt tgtcaagacc gacctgtccg
     5221 gtgccctgaa tgaactgcag gacgaggcag cgcggctatc gtggctggcc acgacgggcg
     5281 ttccttgcgc agctgtgctc gacgttgtca ctgaagcggg aagggactgg ctgctattgg
     5341 gcgaagtgcc ggggcaggat ctcctgtcat ctcaccttgc tcctgccgag aaagtatcca
     5401 tcatggctga tgcaatgcgg cggctgcata cgcttgatcc ggctacctgc ccattcgacc
     5461 accaagcgaa acatcgcatc gagcgagcac gtactcggat ggaagccggt cttgtcgatc
     5521 aggatgatct ggacgaagag catcaggggc tcgcgccagc cgaactgttc gccaggctca
     5581 aggcgcgcat gcccgacggc gaggatctcg tcgtgaccca tggcgatgcc tgcttgccga
     5641 atatcatggt ggaaaatggc cgcttttctg gattcatcga ctgtggccgg ctgggtgtgg
     5701 cggaccgcta tcaggacata gcgttggcta cccgtgatat tgctgaagag cttggcggcg
     5761 aatgggctga ccgcttcctc gtgctttacg gtatcgccgc tcccgattcg cagcgcatcg
     5821 ccttctatcg ccttcttgac gagttcttct gagcgggact ctggggttcg aaatgaccga
     5881 ccaagcgacg cccaacctgc catcacgaga tttcgattcc accgccgcct tctatgaaag
     5941 gttgggcttc ggaatcgttt tccgggacgc cggctggatg atcctccagc gcggggatct
     6001 catgctggag ttcttcgccc accccaactt gtttattgca gcttataatg gttacaaata
     6061 aagcaatagc atcacaaatt tcacaaataa agcatttttt tcactgcatt ctagttgtgg
     6121 tttgtccaaa ctcatcaatg tatcttatca tgtctgtata ccgtcgacct ctagctagag
     6181 cttggcgtaa tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc
     6241 acacaacata cgagccggaa gcataaagtg taaagcctgg ggtgcctaat gagtgagcta
     6301 actcacatta attgcgttgc gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca
     6361 gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc
     6421 cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc
     6481 tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat
     6541 gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt
     6601 ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg
     6661 aaacccggca ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc
     6721 tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt
     6781 ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa
     6841 gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta
     6901 tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa
     6961 caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa
     7021 ctacggctac actagaagaa cagtatttgg tatctgcgct ctgctgaagc cagttacctt
     7081 cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt
     7141 ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat
     7201 cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat
     7261 gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa gttttaaatc
     7321 aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa tcagtgaggc
     7381 acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc ccgtcgtgta
     7441 gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga taccgcgaga
     7501 cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa gggccgagcg
     7561 cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt gccgggaagc
     7621 tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg ctacaggcat
     7681 cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc aacgatcaag
     7741 gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat
     7801 cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag cactgcataa
     7861 ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt actcaaccaa
     7921 gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt caatacggga
     7981 taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac gttcttcggg
     8041 gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac ccactcgtgc
     8101 acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag caaaaacagg
     8161 aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa tactcatact
     8221 cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga gcggatacat
     8281 atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc cccgaaaagt
     8341 gccacctgac gtc
//
