LOCUS       Exported                7592 bp ds-DNA     circular SYN 24-APR-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7592)
  AUTHORS   Nico
  TITLE     Direct Submission
  JOURNAL   Exported Wednesday, May 28, 2025 from SnapGene 5.0.8
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..7592
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     enhancer        5..384
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        386..661
                     /label=chicken beta-actin promoter
     intron          662..1674
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1598..1620
                     /label=pCAGGS-F
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1783..1809
                     /label=mIzumo1 F NheI
     misc_feature    1832..3023
                     /label=Izumo1
     CDS             1841..3145
                     /codon_start=1
                     /label=Izumo1 tagged
                     /translation="MGPHFTLLLAALANCLCPGRPCIKCDQFVTDALKTFENTYLNDHL
                     PHDIHKNVMRMVNHEVSSFGVVTSAEDSYLGAVDENTLEQATWSFLKDLKRITDSDLKG
                     ELFIKELLWMLRHQKDIFNNLARQFQKEVLCPNKCGVMSQTLIWCLKCEKQLHICRKSL
                     DCGERHIEVHRSEDLVLDCLLSWHRASKGLTDYSFYRVWENSSETLIAKGKEPYLTKSM
                     VGPEDAGNYRCVLDTINQGHATVIRYDVTVLPPKHSEENQPPNIITQEEHETPVHVTPQ
                     TPPGQEPESELYPELHPELYPELIPTVAQNPEKKMKTRLLILLTLGFVVLVASIIISVL
                     HFRKVSAKLKNASDEVKPTASGSKSDQSLSQQMGLKKASQADFNSDYSGDKSEATENTR
                     TRPSSLEGPRFEGKPIPNPLLGLDSTRTGHHHHHH"
     primer_bind     complement(3035..3051)
                     /label=mIzumo1 Rvs
     CDS             3074..3115
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     primer_bind     complement(3116..3146)
                     /label=6HisStopSacI_anti
     primer_bind     complement(3116..3145)
                     /label=6HisStopXho_anti
     primer_bind     complement(3125..3156)
                     /label=Izumo R SmaI
     primer_bind     complement(3125..3147)
                     /label=6HisStopNheI_anti
     CDS             3125..3142
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     misc_feature    3161..3747
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(3328..3345)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     3555..3574
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             3748..4509
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYMSGGLQKKLEELELD"
     primer_bind     complement(3793..3814)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3793..3809)
                     /label=pEGFP_N
     primer_bind     complement(4054..4073)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     4401..4422
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(4437..4453)
                     /label=EGFP CR
     primer_bind     4458..4510
                     /label=CNES Fwd
     primer_bind     complement(4462..4514)
                     /label=CNES Rvs
     CDS             4474..4506
                     /codon_start=1
                     /label=NES
                     /translation="LQKKLEELELD"
     primer_bind     complement(4476..4496)
                     /label=NES Colony R
     primer_bind     complement(4584..4603)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_site      4649..4704
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal"
     primer_bind     complement(4703..4722)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(5095..5111)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5095..5111)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5108..5130)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5119..5135
                     /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(5143..5173)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5188..5209
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        5268..5463
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      5314..5449
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     5376..5395
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     primer_bind     complement(5399..5410)
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     polyA_signal    5469..5603
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5506..5525)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5560..5579
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5671..5688)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5842..6430)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5922..5941)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(6601..7461)
                     /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     7224..7243
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7462..7566)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ggtcgacatt gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat
       61 agcccatata tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg
      121 cccaacgacc cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata
      181 gggactttcc attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta
      241 catcaagtgt atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc
      301 gcctggcatt atgcccagta catgacctta tgggactttc ctacttggca gtacatctac
      361 gtattagtca tcgctattac catggtcgag gtgagcccca cgttctgctt cactctcccc
      421 atctcccccc cctccccacc cccaattttg tatttattta ttttttaatt attttgtgca
      481 gcgatggggg cggggggggg gggggcgcgc gccaggcggg gcggggcggg gcgaggggcg
      541 gggcggggcg aggcggagag gtgcggcggc agccaatcag agcggcgcgc tccgaaagtt
      601 tccttttatg gcgaggcggc ggcggcggcg gccctataaa aagcgaagcg cgcggcgggc
      661 gggagtcgct gcgttgcctt cgccccgtgc cccgctccgc gccgcctcgc gccgcccgcc
      721 ccggctctga ctgaccgcgt tactcccaca ggtgagcggg cgggacggcc cttctcctcc
      781 gggctgtaat tagcgcttgg tttaatgacg gctcgtttct tttctgtggc tgcgtgaaag
      841 ccttaaaggg ctccgggagg gccctttgtg cgggggggag cggctcgggg ggtgcgtgcg
      901 tgtgtgtgtg cgtggggagc gccgcgtgcg gcccgcgctg cccggcggct gtgagcgctg
      961 cgggcgcggc gcggggcttt gtgcgctccg cgtgtgcgcg aggggagcgc ggccgggggc
     1021 ggtgccccgc ggtgcggggg ggctgcgagg ggaacaaagg ctgcgtgcgg ggtgtgtgcg
     1081 tgggggggtg agcagggggt gtgggcgcgg cggtcgggct gtaacccccc cctgcacccc
     1141 cctccccgag ttgctgagca cggcccggct tcgggtgcgg ggctccgtac ggggcgtggc
     1201 gcggggctcg ccgtgccggg cggggggtgg cggcaggtgg gggtgccggg cggggcgggg
     1261 ccgcctcggg ccggggaggg ctcgggggag gggcgcggcg gcccccggag cgccggcggc
     1321 tgtcgaggcg cggcgagccg cagccattgc cttttatggt aatcgtgcga gagggcgcag
     1381 ggacttcctt tgtcccaaat ctgtgcggag ccgaaatctg ggaggcgccg ccgcaccccc
     1441 tctagcgggc gcggggcgaa gcggtgcggc gccggcagga aggaaatggg cggggagggc
     1501 cttcgtgcgt cgccgcgccg ccgtcccctt ctccctctcc agcctcgggg ctgtccgcgg
     1561 ggggacggct gccttcgggg gggacggggc agggcggggt tcggcttctg gcgtgtgacc
     1621 ggcggctcta gagcctctgc taaccatgtt catgccttct tctttttcct acagctcctg
     1681 ggcaacgtgc tggttattgt gctgtctcat cattttggca aagaattgct cgagagttcg
     1741 aagccttaag tacgcgtagg cgcgccttaa ttaacgttta aacgctagcg aattcggtcg
     1801 actggatccg gtaccgagga gatctgccgc cgcgatcgcc atggggccgc attttacact
     1861 cttgctggca gctcttgcca actgcctgtg tccagggagg ccctgcatca aatgtgacca
     1921 gtttgtgaca gatgcgctaa agactttcga aaacacttac ctgaatgacc acctgccaca
     1981 cgacattcac aaaaatgtaa tgaggatggt gaaccatgaa gtatcgagct tcggcgtagt
     2041 cacttcggct gaggattcct atttgggggc cgtggacgag aacacactgg aacaagcaac
     2101 ctggagtttt ctgaaggatc tgaagcgtat tacagacagt gacttaaaag gagagctctt
     2161 tataaaggaa ctattgtgga tgcttcgtca tcaaaaggac atctttaaca atcttgctag
     2221 acagttccaa aaggaagttc tttgtcccaa caaatgcgga gtgatgtcgc agactttgat
     2281 ctggtgtctt aagtgcgaaa agcagttgca catttgtcgg aaatccctag attgtggaga
     2341 gcgccacata gaggtacatc gctcggaaga cctggtcctg gactgtctgc tcagttggca
     2401 tcgtgcttct aagggactta cagattacag tttttacagg gtttgggaga acagttctga
     2461 gaccttgatt gccaagggga aagaaccata tctgaccaag tcgatggtgg gtccagagga
     2521 tgctggcaac taccgctgtg tgctagatac catcaaccaa ggtcatgcca ccgtcatccg
     2581 ctacgatgtc acagtattgc ccccaaagca ttcagaggaa aaccaaccac cgaacatcat
     2641 aacccaagag gagcacgaga ctcctgtcca cgtgactcca cagacaccac cggggcagga
     2701 gccagagtcg gagctgtacc cggagctgca cccagagttg tacccggagc tcatacctac
     2761 agtcgctcaa aatccagaga aaaaaatgaa aactcgcctc ttaatactgt tgacccttgg
     2821 ctttgtggtt cttgtggcca gcatcattat ctcggtactt cactttagga aagtcagcgc
     2881 taaattgaag aatgcaagtg acgaagtcaa acccaccgcg tcaggatcca agtcagatca
     2941 gagtttgtcc caacagatgg gattgaagaa ggcctctcag gcagatttta actctgacta
     3001 ctctggagat aagagcgagg caacagaaaa cacgcgtacg cggccctcga gtctagaggg
     3061 cccgcggttc gaaggtaagc ctatccctaa ccctctcctc ggtctcgatt ctacgcgtac
     3121 cggtcatcat caccatcacc attgagcccc gggggatccg cccctctccc tccccccccc
     3181 ctaacgttac tggccgaagc cgcttggaat aaggccggtg tgcgtttgtc tatatgttat
     3241 tttccaccat attgccgtct tttggcaatg tgagggcccg gaaacctggc cctgtcttct
     3301 tgacgagcat tcctaggggt ctttcccctc tcgccaaagg aatgcaaggt ctgttgaatg
     3361 tcgtgaagga agcagttcct ctggaagctt cttgaagaca aacaacgtct gtagcgaccc
     3421 tttgcaggca gcggaacccc ccacctggcg acaggtgcct ctgcggccaa aagccacgtg
     3481 tataagatac acctgcaaag gcggcacaac cccagtgcca cgttgtgagt tggatagttg
     3541 tggaaagagt caaatggctc tcctcaagcg tattcaacaa ggggctgaag gatgcccaga
     3601 aggtacccca ttgtatggga tctgatctgg ggcctcggta cacatgcttt acatgtgttt
     3661 agtcgaggtt aaaaaaacgt ctaggccccc cgaaccacgg ggacgtggtt ttcctttgaa
     3721 aaacacgatg ataatatggc cacaaccatg gtgagcaagg gcgaggagct gttcaccggg
     3781 gtggtgccca tcctggtcga gctggacggc gacgtaaacg gccacaagtt cagcgtgtcc
     3841 ggcgagggcg agggcgatgc cacctacggc aagctgaccc tgaagttcat ctgcaccacc
     3901 ggcaagctgc ccgtgccctg gcccaccctc gtgaccaccc tgacctacgg cgtgcagtgc
     3961 ttcagccgct accccgacca catgaagcag cacgacttct tcaagtccgc catgcccgaa
     4021 ggctacgtcc aggagcgcac catcttcttc aaggacgacg gcaactacaa gacccgcgcc
     4081 gaggtgaagt tcgagggcga caccctggtg aaccgcatcg agctgaaggg catcgacttc
     4141 aaggaggacg gcaacatcct ggggcacaag ctggagtaca actacaacag ccacaacgtc
     4201 tatatcatgg ccgacaagca gaagaacggc atcaaggtga acttcaagat ccgccacaac
     4261 atcgaggacg gcagcgtgca gctcgccgac cactaccagc agaacacccc catcggcgac
     4321 ggccccgtgc tgctgcccga caaccactac ctgagcaccc agtccgccct gagcaaagac
     4381 cccaacgaga agcgcgatca catggtcctg ctggagttcg tgaccgccgc cgggatcact
     4441 ctcggcatgg acgagctgta catgagcggc ggcctgcaga agaagctgga ggagctggag
     4501 ctggactaat gtacaagtaa agcggccggc cgccagcaca gtggtcgatc gaccaatgcc
     4561 ctggctcaca aataccactg agatcttttt ccctctgcca aaaattatgg ggacatcatg
     4621 aagccccttg agcatctgac ttctggctaa taaaggaaat ttattttcat tgcaatagtg
     4681 tgttggaatt ttttgtgtct ctcactcgga aggacatatg ggagggcaaa tcatttaaaa
     4741 catcagaatg agtatttggt ttagagtttg gcaacatatg cccatatgct ggctgccatg
     4801 aacaaaggtt ggctataaag aggtcatcag tatatgaaac agccccctgc tgtccattcc
     4861 ttattccata gaaaagcctt gacttgaggt tagatttttt ttatattttg ttttgtgtta
     4921 tttttttctt taacatccct aaaattttcc ttacatgttt tactagccag atttttcctc
     4981 ctctcctgac tactcccagt catagctgtc cctcttctct tatggagatc cctcgacctg
     5041 caccgtcgac cagctggtcg acggtgcacc gtcgaccagc ttggcgtaat catggtcata
     5101 gctgtttcct gtgtgaaatt gttatccgct cacaattcca cacaacatac gagccggaag
     5161 cataaagtgt aaagcctggg gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg
     5221 ctcactgccc gctttccagt cgggaaacct gtcgtgccag cggatccgca tctcaattag
     5281 tcagcaacca tagtcccgcc cctaactccg cccatcccgc ccctaactcc gcccagttcc
     5341 gcccattctc cgccccatgg ctgactaatt ttttttattt atgcagaggc cgaggccgcc
     5401 tcggcctctg agctattcca gaagtagtga ggaggctttt ttggaggcct aggcttttgc
     5461 aaaaagctaa cttgtttatt gcagcttata atggttacaa ataaagcaat agcatcacaa
     5521 atttcacaaa taaagcattt ttttcactgc attctagttg tggtttgtcc aaactcatca
     5581 atgtatctta tcatgtctgg atccgctgca ttaatgaatc ggccaacgcg cggggagagg
     5641 cggtttgcgt attgggcgct cttccgcttc ctcgctcact gactcgctgc gctcggtcgt
     5701 tcggctgcgg cgagcggtat cagctcactc aaaggcggta atacggttat ccacagaatc
     5761 aggggataac gcaggaaaga acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa
     5821 aaaggccgcg ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa
     5881 tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc
     5941 ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc
     6001 cgcctttctc ccttcgggaa gcgtggcgct ttctcaatgc tcacgctgta ggtatctcag
     6061 ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga
     6121 ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc
     6181 gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac
     6241 agagttcttg aagtggtggc ctaactacgg ctacactaga aggacagtat ttggtatctg
     6301 cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca
     6361 aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa
     6421 aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa
     6481 ctcacgttaa gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt
     6541 aaattaaaaa tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag
     6601 ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat
     6661 agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc
     6721 cagtgctgca atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa
     6781 ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca
     6841 gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa
     6901 cgttgttgcc attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt
     6961 cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc
     7021 ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact
     7081 catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc
     7141 tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg
     7201 ctcttgcccg gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct
     7261 catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc
     7321 cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag
     7381 cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac
     7441 acggaaatgt tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg
     7501 ttattgtctc atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt
     7561 tccgcgcaca tttccccgaa aagtgccacc tg
//
