LOCUS       sequence-205794-        9868 bp    DNA     linear   UNA 15-DEC-2025
DEFINITION  Mammalian expression plasmid for codon optimized hNET with internal 
            MYC tag inserted in ECL2.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      natural DNA sequence
  ORGANISM  unspecified
REFERENCE   1  (bases 1 to 9868)
  AUTHORS   Young HJ, Chan M, Selvam B, Szymanski SK, Shukla D, Procko E
  TITLE     Deep Mutagenesis of a Transporter for Uptake of a Non-Native 
            Substrate Identifies Conformationally Dynamic Regions
  JOURNAL   bioRxiv 2021.04.19.440442
REFERENCE   2  (bases 1 to 9868)
  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: "bioRxiv 
            2021.04.19.440442"
COMMENT     Sequence Label: pCEP4-NET-myc
FEATURES             Location/Qualifiers
     source          1..9868
                     /mol_type="genomic DNA"
                     /organism="unspecified"
     primer_bind     complement(755..773)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        841..945
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(946..1014,1015..1806)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(1164..1183)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1977..2565
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2466..2485
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        2996..3141
                     /label=HSV TK promoter
                     /note="herpes simplex virus thymidine kinase promoter"
     CDS             3184..4221
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPDRE
                     MGEAN"
     primer_bind     complement(4219..4238)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4263..4311
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     enhancer        4965..5344
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        5345..5548
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     5499..5519
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     5545..5569
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             6218..6247
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     polyA_signal    7557..7691
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(7581..7600)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     7635..7654
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(7834..7856)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
ORIGIN
        1 cctcctgctc ctgcccctcc tgctcctgcc cctcccgctc ctgctcctgc tcctgttcca
       61 ccgtgggtcc ctttgcagcc aatgcaactt ggacgttttt ggggtctccg gacaccatct
      121 ctatgtcttg gccctgatcc tgagccgccc ggggctcctg gtcttccgcc tcctcgtcct
      181 cgtcctcttc cccgtcctcg tccatggtta tcaccccctc ttctttgagg tccactgccg
      241 ccggagcctt ctggtccaga tgtgtctccc ttctctccta ggccatttcc aggtcctgta
      301 cctggcccct cgtcagacat gattcacact aaaagagatc aatagacatc tttattagac
      361 gacgctcagt gaatacaggg agtgcagact cctgccccct ccaacagccc ccccaccctc
      421 atccccttca tggtcgctgt cagacagatc caggtctgaa aattccccat cctccgaacc
      481 atcctcgtcc tcatcaccaa ttactcgcag cccggaaaac tcccgctgaa catcctcaag
      541 atttgcgtcc tgagcctcaa gccaggcctc aaattcctcg tccccctttt tgctggacgg
      601 tagggatggg gattctcggg acccctcctc ttcctcttca aggtcaccag acagagatgc
      661 tactggggca acggaagaaa agctgggtgc ggcctgtgag gatcagctta tcgatgataa
      721 gctgtcaaac atgagaattc ttgaagacga aagggcctcg tgatacgcct atttttatag
      781 gttaatgtca tgataataat ggtttcttag acgtcaggtg gcacttttcg gggaaatgtg
      841 cgcggaaccc ctatttgttt atttttctaa atacattcaa atatgtatcc gctcatgaga
      901 caataaccct gataaatgct tcaataatat tgaaaaagga agagtatgag tattcaacat
      961 ttccgtgtcg cccttattcc cttttttgcg gcattttgcc ttcctgtttt tgctcaccca
     1021 gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt gggttacatc
     1081 gaactggatc tcaacagcgg taagatcctt gagagttttc gccccgaaga acgttttcca
     1141 atgatgagca cttttaaagt tctgctatgt ggcgcggtat tatcccgtgt tgacgccggg
     1201 caagagcaac tcggtcgccg catacactat tctcagaatg acttggttga gtactcacca
     1261 gtcacagaaa agcatcttac ggatggcatg acagtaagag aattatgcag tgctgccata
     1321 accatgagtg ataacactgc ggccaactta cttctgacaa cgatcggagg accgaaggag
     1381 ctaaccgctt ttttgcacaa catgggggat catgtaactc gccttgatcg ttgggaaccg
     1441 gagctgaatg aagccatacc aaacgacgag cgtgacacca cgatgcctgc agcaatggca
     1501 acaacgttgc gcaaactatt aactggcgaa ctacttactc tagcttcccg gcaacaatta
     1561 atagactgga tggaggcgga taaagttgca ggaccacttc tgcgctcggc ccttccggct
     1621 ggctggttta ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca
     1681 gcactggggc cagatggtaa gccctcccgt atcgtagtta tctacacgac ggggagtcag
     1741 gcaactatgg atgaacgaaa tagacagatc gctgagatag gtgcctcact gattaagcat
     1801 tggtaactgt cagaccaagt ttactcatat atactttaga ttgatttaaa acttcatttt
     1861 taatttaaaa ggatctaggt gaagatcctt tttgataatc tcatgaccaa aatcccttaa
     1921 cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga
     1981 gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg
     2041 gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc
     2101 agagcgcaga taccaaatac tgtccttcta gtgtagccgt agttaggcca ccacttcaag
     2161 aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc
     2221 agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg
     2281 cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac
     2341 accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga
     2401 aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt
     2461 ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag
     2521 cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg
     2581 gcctttttac ggttcctggc cttttgctgg ccttgaagct gtccctgatg gtcgtcatct
     2641 acctgcctgg acagcatggc ctgcaacgcg ggcatcccga tgccgccgga agcgagaaga
     2701 atcataatgg ggaaggccat ccagcctcgc gtcgcgaacg ccagcaagac gtagcccagc
     2761 gcgtcggccc cgagatgcgc cgcgtgcggc tgctggagat ggcggacgcg atggatatgt
     2821 tctgccaagg gttggtttgc gcattcacag ttctccgcaa gaattgattg gctccaattc
     2881 ttggagtggt gaatccgtta gcgaggtgcc gccctgcttc atccccgtgg cccgttgctc
     2941 gcgtttgctg gcggtgtccc cggaagaaat atatttgcat gtctttagtt ctatgatgac
     3001 acaaaccccg cccagcgtct tgtcattggc gaattcgaac acgcagatgc agtcggggcg
     3061 gcgcggtccg aggtccactt cgcatattaa ggtgacgcgt gtggcctcga acaccgagcg
     3121 accctgcagc gacccgctta acagcgtcaa cagcgtgccg cagatcccgg ggggcaatga
     3181 gatatgaaaa agcctgaact caccgcgacg tctgtcgaga agtttctgat cgaaaagttc
     3241 gacagcgtct ccgacctgat gcagctctcg gagggcgaag aatctcgtgc tttcagcttc
     3301 gatgtaggag ggcgtggata tgtcctgcgg gtaaatagct gcgccgatgg tttctacaaa
     3361 gatcgttatg tttatcggca ctttgcatcg gccgcgctcc cgattccgga agtgcttgac
     3421 attggggaat tcagcgagag cctgacctat tgcatctccc gccgtgcaca gggtgtcacg
     3481 ttgcaagacc tgcctgaaac cgaactgccc gctgttctgc agccggtcgc ggaggccatg
     3541 gatgcgatcg ctgcggccga tcttagccag acgagcgggt tcggcccatt cggaccgcaa
     3601 ggaatcggtc aatacactac atggcgtgat ttcatatgcg cgattgctga tccccatgtg
     3661 tatcactggc aaactgtgat ggacgacacc gtcagtgcgt ccgtcgcgca ggctctcgat
     3721 gagctgatgc tttgggccga ggactgcccc gaagtccggc acctcgtgca cgcggatttc
     3781 ggctccaaca atgtcctgac ggacaatggc cgcataacag cggtcattga ctggagcgag
     3841 gcgatgttcg gggattccca atacgaggtc gccaacatct tcttctggag gccgtggttg
     3901 gcttgtatgg agcagcagac gcgctacttc gagcggaggc atccggagct tgcaggatcg
     3961 ccgcggctcc gggcgtatat gctccgcatt ggtcttgacc aactctatca gagcttggtt
     4021 gacggcaatt tcgatgatgc agcttgggcg cagggtcgat gcgacgcaat cgtccgatcc
     4081 ggagccggga ctgtcgggcg tacacaaatc gcccgcagaa gcgcggccgt ctggaccgat
     4141 ggctgtgtag aagtactcgc cgatagtgga aaccgacgcc ccagcactcg tccggatcgg
     4201 gagatggggg aggctaactg aaacacggaa ggagacaata ccggaaggaa cccgcgctat
     4261 gacggcaata aaaagacaga ataaaacgca cgggtgttgg gtcgtttgtt cataaacgcg
     4321 gggttcggtc ccagggctgg cactctgtcg ataccccacc gagaccccat tggggccaat
     4381 acgcccgcgt ttcttccttt tccccacccc accccccaag ttcgggtgaa ggcccagggc
     4441 tcgcagccaa cgtcggggcg gcaggccctg ccatagccac tggccccgtg ggttagggac
     4501 ggggtccccc atggggaatg gtttatggtt cgtgggggtt attattttgg gcgttgcgtg
     4561 gggtcaggtc cacgactgga ctgagcagac agacccatgg tttttggatg gcctgggcat
     4621 ggaccgcatg tactggcgcg acacgaacac cgggcgtctg tggctgccaa acacccccga
     4681 cccccaaaaa ccaccgcgcg gatttctggc gtgccaagct agtcgaccaa ttctcatgtt
     4741 tgacagctta tcatcgcaga tccgggcaac gttgttgcca ttgctgcagg cgcagaactg
     4801 gtaggtatgg aagatctata cattgaatca atattggcaa ttagccatat tagtcattgg
     4861 ttatatagca taaatcaata ttggctattg gccattgcat acgttgtatc tatatcataa
     4921 tatgtacatt tatattggct catgtccaat atgaccgcca tgttgacatt gattattgac
     4981 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
     5041 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
     5101 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
     5161 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
     5221 aagtccgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
     5281 catgacctta cgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
     5341 catggtgatg cggttttggc agtacaccaa tgggcgtgga tagcggtttg actcacgggg
     5401 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
     5461 ggactttcca aaatgtcgta ataaccccgc cccgttgacg caaatgggcg gtaggcgtgt
     5521 acggtgggag gtctatataa gcagagctcg tttagtgaac cgtcagatct ctagaagctg
     5581 ggtaccagct gctagcgcca ccatgttgct ggctcgcatg aacccccagg tgcagcccga
     5641 gaataacggc gctgataccg gtcccgagca gcccctgagg gcaaggaaaa ccgccgagct
     5701 gttggtggta aaggagagga atggcgtaca gtgcttgctc gctcccaggg acggagacgc
     5761 ccaaccccga gagacgtggg gcaagaagat cgactttctg ttgtctgtcg tgggcttcgc
     5821 ggtggatctt gccaatgtgt ggaggttccc ttacctgtgt tacaagaacg gcggaggagc
     5881 cttcttgatt ccttacaccc tgtttctgat aatcgccgga atgccactgt tctacatgga
     5941 gctggcgctg ggtcagtaca atagggaggg agcggcgacc gtatggaaaa tttgcccctt
     6001 cttcaagggc gtcggctacg ccgtgatcct gattgctctc tacgtcgggt tctactacaa
     6061 cgtgatcatc gcctggagtc tgtactacct gttcagctct tttacgctca acttgccatg
     6121 gacagactgc ggacacacgt ggaactcacc caactgcacg gaccccaaac tccttaacgg
     6181 aagcgtgctg ggcagctccg ggggaagcgg aggcggcgag cagaaactga taagcgagga
     6241 ggacctcggt tcccctggcg gatctagtgg taaccatacc aagtatagca agtacaagtt
     6301 tacccccgct gcagagttct acgagagggg cgtcctgcat ctccatgagt ctagcggcat
     6361 acatgacatc ggactccccc aatggcagtt gctcctgtgc ttgatggtcg tagtgatcgt
     6421 gctgtacttt tctctctgga aaggagtcaa gactagtggc aaggtggttt ggatcaccgc
     6481 caccctgccg tattttgtgc ttttcgttct ccttgtgcat ggcgtgaccc ttcccggtgc
     6541 atcaaacgga ataaatgcct atctgcacat agacttctac agacttaagg aggcaacagt
     6601 gtggatagac gccgcaaccc agatattctt cagccttgga gccggttttg gcgttctgat
     6661 cgcgttcgcc tcttacaaca agttcgacaa caactgctac agggatgctc ttctcacctc
     6721 cagcataaac tgtatcacga gcttcgtgag cggatttgcc atcttctcta tcctgggcta
     6781 tatggcgcat gagcataagg ttaacatcga ggacgtggct accgagggtg caggtcttgt
     6841 gttcatcctg tacccagagg caattagcac cctgagcggc agcacctttt gggcggttgt
     6901 attctttgta atgttgttgg ctttgggact cgacagcagt atgggtggaa tggaagcagt
     6961 gattaccggc ctggccgacg acttccaggt tctgaagagg cataggaagc tgttcacgtt
     7021 cggtgttacc ttctcaacgt ttctcttggc tctgttctgc atcaccaaag gtggcatata
     7081 cgtgctgacc ttgctggata cctttgcagc cggcacctct atactgtttg ccgtactgat
     7141 ggaggctatc ggtgtgagct ggttttacgg ggtggaccga ttctctaacg acatccaaca
     7201 aatgatgggc tttcgacccg gtctctactg gcgactgtgc tggaaatttg tgtcacccgc
     7261 cttccttctt tttgtagttg tggtaagcat aatcaacttc aagccgctga cgtacgacga
     7321 ttacatattc cccccctggg cgaattgggt gggctggggt atcgcgctta gcagcatggt
     7381 gcttgttccc atctatgtca tctataagtt cctgagcacc cagggctcac tgtgggaaag
     7441 gttggcctac ggcataacgc ccgaaaacga gcatcatctg gtagcccaaa gggacatcag
     7501 gcagttccag ctgcaacact ggctggcgat ctaatagctc gaggccggca aggccggatc
     7561 cagacatgat aagatacatt gatgagtttg gacaaaccac aactagaatg cagtgaaaaa
     7621 aatgctttat ttgtgaaatt tgtgatgcta ttgctttatt tgtaaccatt ataagctgca
     7681 ataaacaagt taacaacaac aattgcattc attttatgtt tcaggttcag ggggaggtgt
     7741 gggaggtttt ttaaagcaag taaaacctct acaaatgtgg tatggctgat tatgatccgg
     7801 ctgcctcgcg cgtttcggtg atgacggtga aaacctctga cacatgcagc tcccggagac
     7861 ggtcacagct tgtctgtaag cggatgccgg gagcagacaa gcccgtcagg gcgcgtcagc
     7921 gggtgttggc gggtgtcggg gcgcagccat gaggtcgact ctagaggatc gatccccgcc
     7981 ccggacgaac taaacctgac tacgacatct ctgccccttc ttcgcggggc agtgcatgta
     8041 atcccttcag ttggttggta caacttgcca actgggccct gttccacatg tgacacgggg
     8101 ggggaccaaa cacaaagggg ttctctgact gtagttgaca tccttataaa tggatgtgca
     8161 catttgccaa cactgagtgg ctttcatcct ggagcagact ttgcagtctg tggactgcaa
     8221 cacaacattg cctttatgtg taactcttgg ctgaagctct tacaccaatg ctgggggaca
     8281 tgtacctccc aggggcccag gaagactacg ggaggctaca ccaacgtcaa tcagaggggc
     8341 ctgtgtagct accgataagc ggaccctcaa gagggcatta gcaatagtgt ttataaggcc
     8401 cccttgttaa ccctaaacgg gtagcatatg cttcccgggt agtagtatat actatccaga
     8461 ctaaccctaa ttcaatagca tatgttaccc aacgggaagc atatgctatc gaattagggt
     8521 tagtaaaagg gtcctaagga acagcgatat ctcccacccc atgagctgtc acggttttat
     8581 ttacatgggg tcaggattcc acgagggtag tgaaccattt tagtcacaag ggcagtggct
     8641 gaagatcaag gagcgggcag tgaactctcc tgaatcttcg cctgcttctt cattctcctt
     8701 cgtttagcta atagaataac tgctgagttg tgaacagtaa ggtgtatgtg aggtgctcga
     8761 aaacaaggtt tcaggtgacg cccccagaat aaaatttgga cggggggttc agtggtggca
     8821 ttgtgctatg acaccaatat aaccctcaca aaccccttgg gcaataaata ctagtgtagg
     8881 aatgaaacat tctgaatatc tttaacaata gaaatccatg gggtggggac aagccgtaaa
     8941 gactggatgt ccatctcaca cgaatttatg gctatgggca acacataatc ctagtgcaat
     9001 atgatactgg ggttattaag atgtgtccca ggcagggacc aagacaggtg aaccatgttg
     9061 ttacactcta tttgtaacaa ggggaaagag agtggacgcc gacagcagcg gactccactg
     9121 gttgtctcta acacccccga aaattaaacg gggctccacg ccaatggggc ccataaacaa
     9181 agacaagtgg ccactctttt ttttgaaatt gtggagtggg ggcacgcgtc agcccccaca
     9241 cgccgccctg cggttttgga ctgtaaaata agggtgtaat aacttggctg attgtaaccc
     9301 cgctaaccac tgcggtcaaa ccacttgccc acaaaaccac taatggcacc ccggggaata
     9361 cctgcataag taggtgggcg ggccaagata ggggcgcgat tgctgcgatc tggaggacaa
     9421 attacacaca cttgcgcctg agcgccaagc acagggttgt tggtcctcat attcacgagg
     9481 tcgctgagag cacggtgggc taatgttgcc atgggtagca tatactaccc aaatatctgg
     9541 atagcatatg ctatcctaat ctatatctgg gtagcatagg ctatcctaat ctatatctgg
     9601 gtagcatatg ctatcctaat ctatatctgg gtagtatatg ctatcctaat ttatatctgg
     9661 gtagcatagg ctatcctaat ctatatctgg gtagcatatg ctatcctaat ctatatctgg
     9721 gtagtatatg ctatcctaat ctgtatccgg gtagcatatg ctatcctaat agagattagg
     9781 gtagtatatg ctatcctaat ttatatctgg gtagcatata ctacccaaat atctggatag
     9841 catatgctat cctaatctat atctgggt
//
