LOCUS       sequence-503754-        8574 bp    DNA     circular SYN 26-MAR-2026
DEFINITION  Rat CD4 protein expression in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8574)
  AUTHORS   Nettleship JE, Rada H, Owens RJ
  TITLE     Overview of a High-Throughput Pipeline for Streamlining the 
            Production of Recombinant Proteins.
  JOURNAL   Methods Mol Biol. 2019;2025:33-49. doi: 10.1007/978-1-4939-9624-7_2.
  PUBMED    31267447
REFERENCE   2  (bases 1 to 8574)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Mar 26, 2026 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Methods Mol
            Biol."; date: "2019"; volume: "2025:33-49. doi"; pages: " 
            10.1007/978-1-4939-9624-7_2"
COMMENT     Sequence Label: pOPINTTGneoCD4
FEATURES             Location/Qualifiers
     source          1..8574
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        154..533
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        534..737
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     688..708
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     734..758
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     primer_bind     1239..1258
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     protein_bind    1450..1471
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(1486..1491,1492..1509,1510..1516)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1524..1540
                     /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)."
     primer_bind     1548..1564
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             1560..1733
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGE"
     primer_bind     complement(1580..1597)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(1580..1596)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1589..1611)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     CDS             1753..1776
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             2329..2346
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     polyA_signal    2421..2476
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(2475..2494)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(4712..4730)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4798..4902
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4903..4971,4972..5763)
                     /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(5121..5140)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5934..6522
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6423..6442
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6676..6693
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    6810..6831
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(6846..6851,6852..6869,6870..6876)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6884..6900
                     /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)."
     primer_bind     6908..6924
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6962..6982)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        6967..7296
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      7147..7282
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     7209..7228
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             7363..8157
                     /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(7417..7436)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     8027..8046
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    8333..8454
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(8370..8389)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     8424..8443
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
ORIGIN
        1 agatctatac attgaatcaa tattggcaat tagccatatt agtcattggt tatatagcat
       61 aaatcaatat tggctattgg ccattgcata cgttgtatct atatcataat atgtacattt
      121 atattggctc atgtccaata tgaccgccat gttgacattg attattgact agttattaat
      181 agtaatcaat tacggggtca ttagttcata gcccatatat ggagttccgc gttacataac
      241 ttacggtaaa tggcccgcct ggctgaccgc ccaacgaccc ccgcccattg acgtcaataa
      301 tgacgtatgt tcccatagta acgccaatag ggactttcca ttgacgtcaa tgggtggagt
      361 atttacggta aactgcccac ttggcagtac atcaagtgta tcatatgcca agtccgcccc
      421 ctattgacgt caatgacggt aaatggcccg cctggcatta tgcccagtac atgaccttac
      481 gggactttcc tacttggcag tacatctacg tattagtcat cgctattacc atggtgatgc
      541 ggttttggca gtacaccaat gggcgtggat agcggtttga ctcacgggga tttccaagtc
      601 tccaccccat tgacgtcaat gggagtttgt tttggcacca aaatcaacgg gactttccaa
      661 aatgtcgtaa taaccccgcc ccgttgacgc aaatgggcgg taggcgtgta cggtgggagg
      721 tctatataag cagagctcgt ttagtgaacc gtcagatcct cactctcttc cgcatcgctg
      781 tctgcgaggg ccagctgttg ggctcgcggt tgaggacaaa ctcttcgcgg tctttccagt
      841 actcttggat cggaaacccg tcggcctccg aacggtactc cgccaccgag ggacctgagc
      901 gagtccgcat cgaccggatc ggaaaacctc tcgagaaagg cgtctaacca gtcacagtcg
      961 caaggtaggc tgagcaccgt ggcgggcggc agcgggtggc ggtcggggtt gtttctggcg
     1021 gaggtgctgc tgatgatgta attaaagtag gcggtcttga gacggcggat ggtcgaggtg
     1081 aggtgtggca ggcttgagat ccagctgttg gggtgagtac tccctctcaa aagcgggcat
     1141 tacttctgcg ctaagattgt cagtttccaa aaacgaggag gatttgatat tcacctggcc
     1201 cgatctggcc atacacttga gtgacaatga catccacttt gcctttctct ccacaggtgt
     1261 ccactcccag gtccaagttt atactgaatt cttacaatca aaggagatat accatgggga
     1321 tccttcccag ccctgggatg cctgcgctgc tctccctcgt gagccttctc tccgtgctgc
     1381 tgatgggttg cgtagctgaa accggtaccg tcgaaccgac tggaaagcgg gcagtgagcg
     1441 caacgcaatt aatgtgagtt agctcactca ttaggcaccc caggctttac actttatgct
     1501 tccggctcgt atgttgtgtg gaattgtgag cggataacaa tttcacacag gaaacagcta
     1561 tgaccatgat tacggattca ctggccgtcg ttttacaacg tcgtgactgg gaaaaccctg
     1621 gcgttaccca acttaatcgc cttgcagcac atcccccttt cgccagctgg cgtaatagcg
     1681 aagaggcccg caccgatcgc ccttcccaac agttgcgcag cctgaatggc gaatggcgcc
     1741 aagctgttta aactggaagt tctgtttcag ggcccgacat ccatcacggc ctataagagt
     1801 gagggggagt cagcggagtt ctccttccca ctcaaccttg gagaggaaag cctgcaggga
     1861 gagttgagat ggaaggcaga gaaggctcct tcttcccagt cctggatcac cttctcccta
     1921 aagaaccaaa aggtgtctgt gcagaagtct actagcaacc ccaagttcca gctgtccgaa
     1981 acgctcccac tcacccttca gataccccag gtctcccttc agtttgctgg ttctggcaac
     2041 ctgaccctga ctctggacag agggatactg tatcaggaag tgaacctggt ggtgatgaaa
     2101 gtgactcagc ccgacagcaa cactttgacc tgtgaggtga tgggacccac ctcacccaag
     2161 atgagactga tcttgaagca ggagaatcag gaggccaggg tctccaggca ggagaaagtg
     2221 attcaagtgc aggcccctga agcaggggtg tggcaatgtc tactgagtga aggtgaagag
     2281 gtcaagatgg actccaagat ccaggtttta tccaaagggt tgaataaaca tcaccatcac
     2341 catcactaag tgattaacct caggtgcagg cggccgctcg aggccggcaa ggccggatcc
     2401 cccgacctcg acctctggct aataaaggaa atttattttc attgcaatag tgtgttggaa
     2461 ttttttgtgt ctctcactcg gaaggacata tgggagggca aatcatttgg tcgagatccc
     2521 tcggagatct ctagctagag gatcgatccc cgccccggac gaactaaacc tgactacgac
     2581 atctctgccc cttcttcgcg gggcagtgca tgtaatccct tcagttggtt ggtacaactt
     2641 gccaactggg ccctgttcca catgtgacac ggggggggac caaacacaaa ggggttctct
     2701 gactgtagtt gacatcctta taaatggatg tgcacatttg ccaacactga gtggctttca
     2761 tcctggagca gactttgcag tctgtggact gcaacacaac attgccttta tgtgtaactc
     2821 ttggctgaag ctcttacacc aatgctgggg gacatgtacc tcccaggggc ccaggaagac
     2881 tacgggaggc tacaccaacg tcaatcagag gggcctgtgt agctaccgat aagcggaccc
     2941 tcaagagggc attagcaata gtgtttataa ggcccccttg ttaaccctaa acgggtagca
     3001 tatgcttccc gggtagtagt atatactatc cagactaacc ctaattcaat agcatatgtt
     3061 acccaacggg aagcatatgc tatcgaatta gggttagtaa aagggtccta aggaacagcg
     3121 atatctccca ccccatgagc tgtcacggtt ttatttacat ggggtcagga ttccacgagg
     3181 gtagtgaacc attttagtca caagggcagt ggctgaagat caaggagcgg gcagtgaact
     3241 ctcctgaatc ttcgcctgct tcttcattct ccttcgttta gctaatagaa taactgctga
     3301 gttgtgaaca gtaaggtgta tgtgaggtgc tcgaaaacaa ggtttcaggt gacgccccca
     3361 gaataaaatt tggacggggg gttcagtggt ggcattgtgc tatgacacca atataaccct
     3421 cacaaacccc ttgggcaata aatactagtg taggaatgaa acattctgaa tatctttaac
     3481 aatagaaatc catggggtgg ggacaagccg taaagactgg atgtccatct cacacgaatt
     3541 tatggctatg ggcaacacat aatcctagtg caatatgata ctggggttat taagatgtgt
     3601 cccaggcagg gaccaagaca ggtgaaccat gttgttacac tctatttgta acaaggggaa
     3661 agagagtgga cgccgacagc agcggactcc actggttgtc tctaacaccc ccgaaaatta
     3721 aacggggctc cacgccaatg gggcccataa acaaagacaa gtggccactc ttttttttga
     3781 aattgtggag tgggggcacg cgtcagcccc cacacgccgc cctgcggttt tggactgtaa
     3841 aataagggtg taataacttg gctgattgta accccgctaa ccactgcggt caaaccactt
     3901 gcccacaaaa ccactaatgg caccccgggg aatacctgca taagtaggtg ggcgggccaa
     3961 gataggggcg cgattgctgc gatctggagg acaaattaca cacacttgcg cctgagcgcc
     4021 aagcacaggg ttgttggtcc tcatattcac gaggtcgctg agagcacggt gggctaatgt
     4081 tgccatgggt agcatatact acccaaatat ctggatagca tatgctatcc taatctatat
     4141 ctgggtagca taggctatcc taatctatat ctgggtagca tatgctatcc taatctatat
     4201 ctgggtagta tatgctatcc taatttatat ctgggtagca taggctatcc taatctatat
     4261 ctgggtagca tatgctatcc taatctatat ctgggtagta tatgctatcc taatctgtat
     4321 ccgggtagca tatgctatcc taatagagat tagggtagta tatgctatcc taatttatat
     4381 ctgggtagca tatactaccc aaatatctgg atagcatatg ctatcctaat ctatatctgg
     4441 gtagcatatg ctatcctaat ctatatctgg gtagcatagg ctatcctaat ctatatctgg
     4501 gtagcatatg ctatcctaat ctatatctgg gtagtatatg ctatcctaat ttatatctgg
     4561 gtagcatagg ctatcctaat ctatatctgg gtagcatatg ctatcctaat ctatatctgg
     4621 gtagtatatg ctatcctaat ctgtatccgg gtagcatatg ctatcctcat gataagctgt
     4681 caaacatgag aattttcttg aagacgaaag ggcctcgtga tacgcctatt tttataggtt
     4741 aatgtcatga taataatggt ttcttagacg tcaggtggca cttttcgggg aaatgtgcgc
     4801 ggaaccccta tttgtttatt tttctaaata cattcaaata tgtatccgct catgagacaa
     4861 taaccctgat aaatgcttca ataatattga aaaaggaaga gtatgagtat tcaacatttc
     4921 cgtgtcgccc ttattccctt ttttgcggca ttttgccttc ctgtttttgc tcacccagaa
     4981 acgctggtga aagtaaaaga tgctgaagat cagttgggtg cacgagtggg ttacatcgaa
     5041 ctggatctca acagcggtaa gatccttgag agttttcgcc ccgaagaacg ttttccaatg
     5101 atgagcactt ttaaagttct gctatgtggc gcggtattat cccgtgttga cgccgggcaa
     5161 gagcaactcg gtcgccgcat acactattct cagaatgact tggttgagta ctcaccagtc
     5221 acagaaaagc atcttacgga tggcatgaca gtaagagaat tatgcagtgc tgccataacc
     5281 atgagtgata acactgcggc caacttactt ctgacaacga tcggaggacc gaaggagcta
     5341 accgcttttt tgcacaacat gggggatcat gtaactcgcc ttgatcgttg ggaaccggag
     5401 ctgaatgaag ccataccaaa cgacgagcgt gacaccacga tgcctgcagc aatggcaaca
     5461 acgttgcgca aactattaac tggcgaacta cttactctag cttcccggca acaattaata
     5521 gactggatgg aggcggataa agttgcagga ccacttctgc gctcggccct tccggctggc
     5581 tggtttattg ctgataaatc tggagccggt gagcgtgggt ctcgcggtat cattgcagca
     5641 ctggggccag atggtaagcc ctcccgtatc gtagttatct acacgacggg gagtcaggca
     5701 actatggatg aacgaaatag acagatcgct gagataggtg cctcactgat taagcattgg
     5761 taactgtcag accaagttta ctcatatata ctttagattg atttaaaact tcatttttaa
     5821 tttaaaagga tctaggtgaa gatccttttt gataatctca tgaccaaaat cccttaacgt
     5881 gagttttcgt tccactgagc gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat
     5941 cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct accagcggtg
     6001 gtttgtttgc cggatcaaga gctaccaact ctttttccga aggtaactgg cttcagcaga
     6061 gcgcagatac caaatactgt tcttctagtg tagccgtagt taggccacca cttcaagaac
     6121 tctgtagcac cgcctacata cctcgctctg ctaatcctgt taccagtggc tgctgccagt
     6181 ggcgataagt cgtgtcttac cgggttggac tcaagacgat agttaccgga taaggcgcag
     6241 cggtcgggct gaacgggggg ttcgtgcaca cagcccagct tggagcgaac gacctacacc
     6301 gaactgagat acctacagcg tgagctatga gaaagcgcca cgcttcccga agggagaaag
     6361 gcggacaggt atccggtaag cggcagggtc ggaacaggag agcgcacgag ggagcttcca
     6421 gggggaaacg cctggtatct ttatagtcct gtcgggtttc gccacctctg acttgagcgt
     6481 cgatttttgt gatgctcgtc aggggggcgg agcctatgga aaaacgccag caacgcggcc
     6541 tttttacggt tcctggcctt ttgctggcct tttgctcaca tgttctttcc tgcgttatcc
     6601 cctgattctg tggataaccg tattaccgcc tttgagtgag ctgataccgc tcgccgcagc
     6661 cgaacgaccg agcgcagcga gtcagtgagc gaggaagcgg aagagcgccc aatacgcaaa
     6721 ccgcctctcc ccgcgcgttg gccgattcat taatgcagct ggcacgacag gtttcccgac
     6781 tggaaagcgg gcagtgagcg caacgcaatt aatgtgagtt agctcactca ttaggcaccc
     6841 caggctttac actttatgct tccggctcgt atgttgtgtg gaattgtgag cggataacaa
     6901 tttcacacag gaaacagcta tgaccatgat tacgccaagc tctagctaga ggtcgacctg
     6961 tggaatgtgt gtcagttagg gtgtggaaag tccccaggct ccccagcagg cagaagtatg
     7021 caaagcatgc atctcaatta gtcagcaacc aggtgtggaa agtccccagg ctccccagca
     7081 ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccatagtccc gcccctaact
     7141 ccgcccatcc cgcccctaac tccgcccagt tccgcccatt ctccgcccca tggctgacta
     7201 atttttttta tttatgcaga ggccgaggcc gcctctgcct ctgagctatt ccagaagtag
     7261 tgaggaggct tttttggagg cctaggcttt tgcaaaaagc tcccgggagc ttgtatatcc
     7321 attttcggat ctgatcaaga gacaggatga ggatcgtttc gcatgattga acaagatgga
     7381 ttgcacgcag gttctccggc cgcttgggtg gagaggctat tcggctatga ctgggcacaa
     7441 cagacaatcg gctgctctga tgccgccgtg ttccggctgt cagcgcaggg gcgcccggtt
     7501 ctttttgtca agaccgacct gtccggtgcc ctgaatgaac tgcaggacga ggcagcgcgg
     7561 ctatcgtggc tggccacgac gggcgttcct tgcgcagctg tgctcgacgt tgtcactgaa
     7621 gcgggaaggg actggctgct attgggcgaa gtgccggggc aggatctcct gtcatctcac
     7681 cttgctcctg ccgagaaagt atccatcatg gctgatgcaa tgcggcggct gcatacgctt
     7741 gatccggcta cctgcccatt cgaccaccaa gcgaaacatc gcatcgagcg agcacgtact
     7801 cggatggaag ccggtcttgt cgatcaggat gatctggacg aagagcatca ggggctcgcg
     7861 ccagccgaac tgttcgccag gctcaaggcg cgcatgcccg acggcgagga tctcgtcgtg
     7921 acccatggcg atgcctgctt gccgaatatc atggtggaaa atggccgctt ttctggattc
     7981 atcgactgtg gccggctggg tgtggcggac cgctatcagg acatagcgtt ggctacccgt
     8041 gatattgctg aagagcttgg cggcgaatgg gctgaccgct tcctcgtgct ttacggtatc
     8101 gccgctcccg attcgcagcg catcgccttc tatcgccttc ttgacgagtt cttctgagcg
     8161 ggactctggg gttcgcgaaa tgaccgacca agcgacgccc aacctgccat cacgagattt
     8221 cgattccacc gccgccttct atgaaaggtt gggcttcgga atcgttttcc gggacgccgg
     8281 ctggatgatc ctccagcgcg gggatctcat gctggagttc ttcgcccacc ccaacttgtt
     8341 tattgcagct tataatggtt acaaataaag caatagcatc acaaatttca caaataaagc
     8401 atttttttca ctgcattcta gttgtggttt gtccaaactc atcaatgtat cttatcatgt
     8461 ctgtataccg tcgacctcta gctagtcgac caattctcat gtttgacagc ttatcatcgc
     8521 agatccgggc aacgttgttg ccattgctgc aggcgcagaa ctggtaggta tgga
//
