LOCUS       sequence-142719-        7122 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses NET1A-V5 in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7122)
  AUTHORS   Schaffer BE, Levin RS, Hertz NT, Maures TJ, Schoof ML, Hollstein PE,
            Benayoun BA, Banko MR, Shaw RJ, Shokat KM, Brunet A
  TITLE     Identification of AMPK Phosphorylation Sites Reveals a Network of 
            Proteins Involved in Cell Invasion and Facilitates Large-Scale 
            Substrate Prediction.
  JOURNAL   Cell Metab. 2015 Nov 3;22(5):907-21. doi: 
            10.1016/j.cmet.2015.09.009. Epub 2015 Oct 8.
  PUBMED    26456332
REFERENCE   2  (bases 1 to 7122)
  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"; doi: 
            "10.1016/j.cmet.2015.09.009"; journalName: "Cell Metab"; date: 
            "2015-11-3- 3"; volume: "22"; issue: "5"; pages: "907-21"
COMMENT     Sequence Label: pcDNA 3.2/V5-DEST NET1A S46A
FEATURES             Location/Qualifiers
     source          1..7122
                     /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"
     protein_bind    911..935
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(2562..2586)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             2612..2653
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     primer_bind     complement(2693..2712)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    2737..2785
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      2987..3415
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3074..3093)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3284..3305
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3424..3444)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3429..3758
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3609..3744
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3671..3690
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3825..4619
                     /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(3879..3898)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4489..4508
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4795..4916
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4832..4851)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4886..4905
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4965..4981)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4965..4981)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4978..5000)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4989..5005
                     /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(5013..5019,5020..5037,5038..5043))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5058..5079
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5196..5213)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5367..5955)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5447..5466)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6126..6917,6918..6986))
                     /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     6749..6768
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6987..7091)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtcgactct 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 atgggtggac 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 gctggctagt
      901 taagctatca acaagtttgt acaaaaaagt tggcatggtg gcacatgatg agactggagg
      961 tctcctacct attaaaagga ccatacgagt cctagatgtc aataaccagt ccttcagaga
     1021 acaagaggag ccaagcaata aaagagttcg acctctggct cgtgtcacgg ccttggcaaa
     1081 tttaatctct cctgtaagaa atggagctgt cagacgtttt ggtcaaacaa tacagtcatt
     1141 tacccttcgt ggtgaccaca gatccccagc ctctgcccag aagttttcta gcaggtcaac
     1201 agtcccaaca cccgccaaga gaaggagcag tgcactgtgg tcagagatgc tggacatcac
     1261 catgaaggag tctctcacca ccagggagat cagacggcag gaggcaatat atgaaatgtc
     1321 ccgaggtgaa caggatttaa ttgaggatct caaacttgca agaaaggcct accatgaccc
     1381 catgttaaag ttgtccatca tgtcagaaga ggaactcaca catatatttg gtgatctgga
     1441 ctcttacata cctctgcatg aagatttgtt gacaagaata ggagaagcaa ccaagcctga
     1501 tggaacagtg gagcagattg gtcacattct cgtgagctgg ttaccgcgct tgaatgccta
     1561 cagaggttac tgtagtaacc agctggcagc caaagctctt cttgatcaaa agaaacagga
     1621 tccaagagtc caagacttcc tccagcgatg tctcgagtct cccttcagtc gaaaactaga
     1681 tctttggagt ttcctagata tccctcgaag tcgcctagtc aaataccctt tactgttaaa
     1741 agaaattctt aaacacactc caaaagagca ccctgatgtt cagcttctgg aggatgctat
     1801 attgataata cagggagtcc tctctgatat caacttgaag aaaggtgaat ccgagtgcca
     1861 gtattacatc gacaagctgg agtacctgga tgaaaagcag agggacccca gaatcgaagc
     1921 gagcaaagtg ctgctgtgcc atggggagct gcggagcaag agtggacata aactttacat
     1981 tttcctgttt caagacatct tggttctgac tcggcccgtc acacggaacg aacggcactc
     2041 ttaccaggtt taccggcagc caatcccagt ccaagagcta gtcctagaag acctgcagga
     2101 tggagatgtg agaatgggag gctcctttcg aggagctttc agtaactcag agaaagctaa
     2161 aaatatcttt agaattcgct tccatgaccc ctctccagcc cagtctcaca ctctgcaagc
     2221 caatgacgtg ttccacaagc agcagtggtt caactgtatt cgagcggcca ttgccccctt
     2281 ccagtcggca ggcagtccac ctgagctgca gggcctgccg gagctgcacg aagagtgtga
     2341 ggggaaccac ccctctgcga ggaaactcac agcccagagg agggcatcca cagtttccag
     2401 tgttactcag gtagaagttg atgaaaacgc ttacagatgt ggctctggca tgcagatggc
     2461 agaggacagc aagagcttaa agacacacca gacacagccc ggcatccgaa gagcgaggga
     2521 caaagccctt tctggtggca aacggaaaga gactttggtg tacccaactt tcttgtacaa
     2581 agtggttgat ctagagggcc cgcggttcga aggtaagcct atccctaacc ctctcctcgg
     2641 tctcgattct acgcgtaccg gttagtaatg agtttaaacg ggggaggcta actgaaacac
     2701 ggaaggagac aataccggaa ggaacccgcg ctatgacggc aataaaaaga cagaataaaa
     2761 cgcacgggtg ttgggtcgtt tgttcataaa cgcggggttc ggtcccaggg ctggcactct
     2821 gtcgataccc caccgagacc ccattggggc caatacgccc gcgtttcttc cttttcccca
     2881 ccccaccccc caagttcggg tgaaggccca gggctcgcag ccaacgtcgg ggcggcaggc
     2941 cctgccatag cagatctgcg cagctggggc tctagggggt atccccacgc gccctgtagc
     3001 ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac acttgccagc
     3061 gccctagcgc ccgctccttt cgctttcttc ccttcctttc tcgccacgtt cgccggcttt
     3121 ccccgtcaag ctctaaatcg gggcatccct ttagggttcc gatttagtgc tttacggcac
     3181 ctcgacccca aaaaacttga ttagggtgat ggttcacgta gtgggccatc gccctgatag
     3241 acggtttttc gccctttgac gttggagtcc acgttcttta atagtggact cttgttccaa
     3301 actggaacaa cactcaaccc tatctcggtc tattcttttg atttataagg gattttgggg
     3361 atttcggcct attggttaaa aaatgagctg atttaacaaa aatttaacgc gaattaattc
     3421 tgtggaatgt gtgtcagtta gggtgtggaa agtccccagg ctccccagca ggcagaagta
     3481 tgcaaagcat gcatctcaat tagtcagcaa ccaggtgtgg aaagtcccca ggctccccag
     3541 caggcagaag tatgcaaagc atgcatctca attagtcagc aaccatagtc ccgcccctaa
     3601 ctccgcccat cccgccccta actccgccca gttccgccca ttctccgccc catggctgac
     3661 taattttttt tatttatgca gaggccgagg ccgcctctgc ctctgagcta ttccagaagt
     3721 agtgaggagg cttttttgga ggcctaggct tttgcaaaaa gctcccggga gcttgtatat
     3781 ccattttcgg atctgatcaa gagacaggat gaggatcgtt tcgcatgatt gaacaagatg
     3841 gattgcacgc aggttctccg gccgcttggg tggagaggct attcggctat gactgggcac
     3901 aacagacaat cggctgctct gatgccgccg tgttccggct gtcagcgcag gggcgcccgg
     3961 ttctttttgt caagaccgac ctgtccggtg ccctgaatga actgcaggac gaggcagcgc
     4021 ggctatcgtg gctggccacg acgggcgttc cttgcgcagc tgtgctcgac gttgtcactg
     4081 aagcgggaag ggactggctg ctattgggcg aagtgccggg gcaggatctc ctgtcatctc
     4141 accttgctcc tgccgagaaa gtatccatca tggctgatgc aatgcggcgg ctgcatacgc
     4201 ttgatccggc tacctgccca ttcgaccacc aagcgaaaca tcgcatcgag cgagcacgta
     4261 ctcggatgga agccggtctt gtcgatcagg atgatctgga cgaagagcat caggggctcg
     4321 cgccagccga actgttcgcc aggctcaagg cgcgcatgcc cgacggcgag gatctcgtcg
     4381 tgacccatgg cgatgcctgc ttgccgaata tcatggtgga aaatggccgc ttttctggat
     4441 tcatcgactg tggccggctg ggtgtggcgg accgctatca ggacatagcg ttggctaccc
     4501 gtgatattgc tgaagagctt ggcggcgaat gggctgaccg cttcctcgtg ctttacggta
     4561 tcgccgctcc cgattcgcag cgcatcgcct tctatcgcct tcttgacgag ttcttctgag
     4621 cgggactctg gggttcgcga aatgaccgac caagcgacgc ccaacctgcc atcacgagat
     4681 ttcgattcca ccgccgcctt ctatgaaagg ttgggcttcg gaatcgtttt ccgggacgcc
     4741 ggctggatga tcctccagcg cggggatctc atgctggagt tcttcgccca ccccaacttg
     4801 tttattgcag cttataatgg ttacaaataa agcaatagca tcacaaattt cacaaataaa
     4861 gcattttttt cactgcattc tagttgtggt ttgtccaaac tcatcaatgt atcttatcat
     4921 gtctgtatac cgtcgacctc tagctagagc ttggcgtaat catggtcata gctgtttcct
     4981 gtgtgaaatt gttatccgct cacaattcca cacaacatac gagccggaag cataaagtgt
     5041 aaagcctggg gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc
     5101 gctttccagt cgggaaacct gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg
     5161 agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg
     5221 gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca
     5281 gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac
     5341 cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac
     5401 aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg
     5461 tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac
     5521 ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc aatgctcacg ctgtaggtat
     5581 ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag
     5641 cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac
     5701 ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt
     5761 gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaaggac agtatttggt
     5821 atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc
     5881 aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga
     5941 aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac
     6001 gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc
     6061 cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct
     6121 gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca
     6181 tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct
     6241 ggccccagtg ctgcaatgat accgcgagac ccacgctcac cggctccaga tttatcagca
     6301 ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc
     6361 atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg
     6421 cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct
     6481 tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa
     6541 aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta
     6601 tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc
     6661 ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg
     6721 agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag aactttaaaa
     6781 gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg
     6841 agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc
     6901 accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg
     6961 gcgacacgga aatgttgaat actcatactc ttcctttttc aatattattg aagcatttat
     7021 cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata
     7081 ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg tc
//
