LOCUS       sequence-223821-        6876 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of the extracellular domain of NGRN with N-terminal FLAG 
            tag and C-terminal hIgG tag..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6876)
  TITLE     Comoletti Lab Plasmids
REFERENCE   2  (bases 1 to 6876)
  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"
COMMENT     Sequence Label: pCMV6-XL4 FLAG-NGRN-Fc
FEATURES             Location/Qualifiers
     source          1..6876
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     151..173
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     165..182
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     166..182
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     enhancer        343..722
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        723..926
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     877..897
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      950..959
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             956..1045
                     /codon_start=1
                     /product="signal sequence from bovine prolactin"
                     /label=prolactin signal sequence
                     /translation="MDSKGSSQKGSRLLLLLVVSNLLLCQGVVS"
     CDS             1046..1069
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             1910..1933
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     polyA_signal    3242..3864
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(3441..3460)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     primer_bind     complement(3888..3908)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3893..4222
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4073..4208
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(4260..4279)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(4261..4279)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4293..4309)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4293..4309)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4306..4328)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4317..4333
                     /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(4341..4347,4348..4365,4366..4371))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4386..4407
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4524..4541)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4695..5283)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4775..4794)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5454..6245,6246..6314))
                     /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     6077..6096
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6315..6419)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      6446..6874
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6533..6552)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6743..6764
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
ORIGIN
        1 aacaaaatat taacgcttac aatttccatt cgccattcag gctgcgcaac tgttgggaag
       61 ggcgatcggt gcgggcctct tcgctattac gccagctggc gaaaggggga tgtgctgcaa
      121 ggcgattaag ttgggtaacg ccagggtttt cccagtcacg acgttgtaaa acgacggcca
      181 gtgccaagct gatctataca ttgaatcaat attggcaatt agccatatta gtcattggtt
      241 atatagcata aatcaatatt ggctattggc cattgcatac gttgtatcta tatcataata
      301 tgtacattta tattggctca tgtccaatat gaccgccatg ttgacattga ttattgacta
      361 gttattaata gtaatcaatt acggggtcat tagttcatag cccatatatg gagttccgcg
      421 ttacataact tacggtaaat ggcccgcctg gctgaccgcc caacgacccc cgcccattga
      481 cgtcaataat gacgtatgtt cccatagtaa cgccaatagg gactttccat tgacgtcaat
      541 gggtggagta tttacggtaa actgcccact tggcagtaca tcaagtgtat catatgccaa
      601 gtccgccccc tattgacgtc aatgacggta aatggcccgc ctggcattat gcccagtaca
      661 tgaccttacg ggactttcct acttggcagt acatctacgt attagtcatc gctattacca
      721 tggtgatgcg gttttggcag tacaccaatg ggcgtggata gcggtttgac tcacggggat
      781 tttcaagtct ccaccccatt gacgtcaatg ggagtttgtt ttggcaccaa aatcaacggg
      841 actttccaaa atgtcgtaat aaccccgccc cgttgacgca aatgggcggt aggcgtgtac
      901 ggtgggaggt ctatataagc agagctcgtt tagtgaaccg tcagatatcg ccaccatgga
      961 cagcaaaggt tcgtcgcaga aagggtcccg cctgctcctg ctgctggtgg tgtcaaatct
     1021 actcttgtgc cagggtgtgg tctccgacta caaagacgat gacgacaagg cggccgctgc
     1081 tgtgaccaga tgcggcttcg ctaccagagg cgtggccggc cctggcccta tcggcagaga
     1141 gcctgatccc gatagcgact gggagcctga ggagagggag ctgcaggagg tggagagcac
     1201 cctgaagaga cagaagcagg ccatcagatt ccagaagatc agaagacaga tggaggcccc
     1261 cggcgccccc cccagaaccc tgacctggga ggccatggag cagatcagat acctgcacga
     1321 ggagttcccc gagagctgga gcgtgcccag actggccgag ggcttcgacg tgagcaccga
     1381 cgtgatcaga agagtgctga agagcaagtt cctgcccacc ctggagcaga agctgaagca
     1441 ggaccagaag gtgctgaaga aggccggcct ggcccacagc ctgcagcacc tgagaggcag
     1501 cggcaatacc agcaaactgc tgcctgccgg ccacagcgtg agcggcagcc tgctgatgcc
     1561 tggccacgag gccagcagca aggaccccaa ccacagcacc gccctgaagg tgatcgagag
     1621 cgacacccac agaaccaaca cccccagaag aagaaagggc agaaacaagg agatccagga
     1681 cctggaggag agcttcgtgc ccgtggccgc ccctctggga caccccagag agctgcagaa
     1741 gtacagcagc gatagcgagt ctcctagagg aacaggcagc ggcgccctgc ccagcggcca
     1801 gaagctggag gagctgaagg ccgaggagcc cgacaacttc agcagcaagg tggtgcagag
     1861 aggcagagag ttcttcgaca gcaacggcaa cttcctgtac agaatcggtc tagaggtgct
     1921 cttccagggg cctgatcccg atcccgaggg tgagtactaa gcttcagcgc tcctgcctgg
     1981 acgcatcccg gctatgcagc cccagtccag ggcagcaagg caggccccgt ctgcctcttc
     2041 acccggaggc ctctgcccgc cccactcatg ctcagggaga gggtcttctg gctttttccc
     2101 caggctctgg gcaggcacag gctaggtgcc cctaacccag gccctgcaca caaaggggca
     2161 ggtgctgggc tcagacctgc caagagccat atccgggagg accctgcccc tgacctaagc
     2221 ccaccccaaa ggccaaactc tccactccct cagctcggac accttctctc ctcccagatt
     2281 ccagtaactc ccaatcttct ctctgcagag cccaaatctt gtgacaaaac tcacacatgc
     2341 ccaccgtgcc caggtaagcc agcccaggcc tcgccctcca gctcaaggcg ggacaggtgc
     2401 cctagagtag cctgcatcca gggacaggcc ccagccgggt gctgacacgt ccacctccat
     2461 ctcttcctca gcacctgaac tcctgggggg accgtcagtc ttcctcttcc ccccaaaacc
     2521 caaggacacc ctcatgatct cccggacccc tgaggtcaca tgcgtggtgg tggacgtgag
     2581 ccacgaagac cctgaggtca agttcaactg gtacgtggac ggcgtggagg tgcataatgc
     2641 caagacaaag ccgcgggagg agcagtacaa cagcacgtac cgtgtggtca gcgtcctcac
     2701 cgtcctgcac caggactggc tgaatggcaa ggagtacaag tgcaaggtct ccaacaaagc
     2761 cctcccagcc cccatcgaga aaaccatctc caaagccaaa ggtgggaccc gtggggtgcg
     2821 agggccacat ggacagaggc cggctcggcc caccctctgc cctgagagtg accgctgtac
     2881 caacctctgt ccctacaggg cagccccgag aaccacaggt gtacaccctg cccccatccc
     2941 gggatgagct gaccaagaac caggtcagcc tgacctgcct ggtcaaaggc ttctatccca
     3001 gcgacatcgc cgtggagtgg gagagcaatg ggcagccgga gaacaactac aagaccacgc
     3061 ctcccgtgct ggactccgac ggctccttct tcctctacag caagctcacc gtggacaaga
     3121 gcaggtggca gcaggggaac gtcttctcat gctccgtgat gcatgaggct ctgcacaacc
     3181 actacacgca gaagagcctc tccctgtctc cgggtaaatg agtgcgacgg ccgcgactct
     3241 agggtggcat ccctgtgacc cctccccagt gcctctcctg gccctggaag ttgccactcc
     3301 agtgcccacc agccttgtcc taataaaatt aagttgcatc attttgtctg actaggtgtc
     3361 cttctataat attatggggt ggaggggggt ggtatggagc aaggggcaag ttgggaagac
     3421 aacctgtagg gcctgcgggg tctattggga accaagctgg agtgcagtgg cacaatcttg
     3481 gctcactgca atctccgcct cctgggttca agcgattctc ctgcctcagc ctcccgagtt
     3541 gttgggattc caggcatgca tgaccaggct cagctaattt ttgttttttt ggtagagacg
     3601 gggtttcacc atattggcca ggctggtctc caactcctaa tctcaggtga tctacccacc
     3661 ttggcctccc aaattgctgg gattacaggc gtgaaccact gctcccttcc ctgtccttct
     3721 gattttaaaa taactatacc agcaggagga cgtccagaca cagcataggc tacctggcca
     3781 tgcccaaccg gtgggacatt tgagttgctt gcttggcact gtcctctcat gcgttgggtc
     3841 cactcagtag atgcctgttg aattgggtac gcggccagct tggctgtgga atgtgtgtca
     3901 gttagggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa gcatgcatct
     3961 caattagtca gcaaccaggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca
     4021 aagcatgcat ctcaattagt cagcaaccat agtcccgccc ctaactccgc ccatcccgcc
     4081 cctaactccg cccagttccg cccattctcc gccccatggc tgactaattt tttttattta
     4141 tgcagaggcc gaggccgcct cggcctctga gctattccag aagtagtgag gaggcttttt
     4201 tggaggccta ggcttttgca aaaagctcct cgaggaactg aaaaaccaga aagttaattc
     4261 cctatagtga gtcgtattaa attcgtaatc atgtcatagc tgtttcctgt gtgaaattgt
     4321 tatccgctca caattccaca caacatacga gccggaagca taaagtgtaa agcctggggt
     4381 gcctaatgag tgagctaact cacattaatt gcgttgcgct cactgcccgc tttccagtcg
     4441 ggaaacctgt cgtgccagct gcattaatga atcggccaac gcgcggggag aggcggtttg
     4501 cgtattgggc gctcttccgc ttcctcgctc actgactcgc tgcgctcggt cgttcggctg
     4561 cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga atcaggggat
     4621 aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc
     4681 gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc
     4741 tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctgga
     4801 agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct gtccgccttt
     4861 ctcccttcgg gaagcgtggc gctttctcat agctcacgct gtaggtatct cagttcggtg
     4921 taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc
     4981 gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt atcgccactg
     5041 gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc tacatagttc
     5101 ttgaagtggt ggcctaacta cggctacact agaagaacag tatttggtat ctgcgctctg
     5161 ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa acaaaccacc
     5221 gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct
     5281 caagaagatc ctttgatctt ttctacgggg tctgacgctc agtggaacga aaactcacgt
     5341 taagggattt tggtcatgag attatcaaaa aggatcttca cctagatcct tttaaattaa
     5401 aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa cttggtctga cagttaccaa
     5461 tgcttaatca gtgaggcacc tatctcagcg atctgtctat ttcgttcatc catagttgcc
     5521 tgactccccg tcgtgtagat aactacgata cgggagggct taccatctgg ccccagtgct
     5581 gcaatgatac cgcgagaccc acgctcaccg gctccagatt tatcagcaat aaaccagcca
     5641 gccggaaggg ccgagcgcag aagtggtcct gcaactttat ccgcctccat ccagtctatt
     5701 aattgttgcc gggaagctag agtaagtagt tcgccagtta atagtttgcg caacgttgtt
     5761 gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc
     5821 ggttcccaac gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa agcggttagc
     5881 tccttcggtc ctccgatcgt tgtcagaagt aagttggccg cagtgttatc actcatggtt
     5941 atggcagcac tgcataattc tcttactgtc atgccatccg taagatgctt ttctgtgact
     6001 ggtgagtact caaccaagtc attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc
     6061 ccggcgtcaa tacgggataa taccgcgcca catagcagaa ctttaaaagt gctcatcatt
     6121 ggaaaacgtt cttcggggcg aaaactctca aggatcttac cgctgttgag atccagttcg
     6181 atgtaaccca ctcgtgcacc caactgatct tcagcatctt ttactttcac cagcgtttct
     6241 gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa
     6301 tgttgaatac tcatactctt cctttttcaa tattattgaa gcatttatca gggttattgt
     6361 ctcatgagcg gatacatatt tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc
     6421 acatttcccc gaaaagtgcc acctgacgcg ccctgtagcg gcgcattaag cgcggcgggt
     6481 gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc cgctcctttc
     6541 gctttcttcc cttcctttct cgccacgttc gccggctttc cccgtcaagc tctaaatcgg
     6601 gggctccctt tagggttccg atttagtgct ttacggcacc tcgaccccaa aaaacttgat
     6661 tagggtgatg gttcacgtag tgggccatcg ccctgataga cggtttttcg ccctttgacg
     6721 ttggagtcca cgttctttaa tagtggactc ttgttccaaa ctggaacaac actcaaccct
     6781 atctcggtct attcttttga tttataaggg attttgccga tttcggccta ttggttaaaa
     6841 aatgagctga tttaacaaaa atttaacgcg aatttt
//
