LOCUS       sequence-223834-        6888 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of the extracellular domain of TMEFF2 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 6888)
  TITLE     Comoletti Lab Plasmids
REFERENCE   2  (bases 1 to 6888)
  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-TMEFF2-Fc
FEATURES             Location/Qualifiers
     source          1..6888
                     /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             1922..1945
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     polyA_signal    3254..3876
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(3453..3472)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     primer_bind     complement(3900..3920)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3905..4234
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4085..4220
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(4272..4291)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(4273..4291)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4305..4321)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4305..4321)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4318..4340)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4329..4345
                     /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(4353..4359,4360..4377,4378..4383))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4398..4419
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4536..4553)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4707..5295)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4787..4806)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5466..6257,6258..6326))
                     /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     6089..6108
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6327..6431)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      6458..6886
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6545..6564)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6755..6776
                     /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 cggccgcttt
     1081 tcccaccagc ctgagcgact gccagacccc caccggctgg aactgcagcg gctacgacga
     1141 cagagagaac gacctgttcc tgtgcgacac caacacctgt aagttcgacg gcgagtgcct
     1201 gagaatcggc gacaccgtga cctgcgtgtg ccagttcaag tgcaacaacg actacgtgcc
     1261 cgtgtgcggc agcaacggcg agagctacca gaacgagtgc tacctgagac aggccgcctg
     1321 taagcagcag agcgagatcc tggtggtgtc tgaaggcagc tgtgccaccg atgccggatc
     1381 tggcagcggc gacggcgtgc atgagggcag cggcgagaca agccagaagg agacaagcac
     1441 atgcgacatc tgccagttcg gcgccgagtg cgacgaggac gccgaggacg tgtggtgcgt
     1501 gtgcaacatc gactgcagcc agaccaactt caaccccctg tgcgccagcg acggcaagag
     1561 ctacgacaac gcctgccaga tcaaggaggc cagctgccag aagcaggaga agatcgaggt
     1621 gatgagcctg ggcagatgcc aggacaacac caccaccacc accaagagcg aggacggcca
     1681 ctacgccaga accgactacg ccgagaacgc caacaagctg gaggagagcg ccagagagca
     1741 ccacatcccc tgccccgagc actacaacgg cttctgcatg cacggcaagt gcgagcacag
     1801 catcaacatg caggagccca gctgcagatg cgacgccggc tacaccggcc agcactgcga
     1861 gaagaaggac tacagcgtgc tgtacgtggt gcccggcccc gtgagattcc agtacgtggg
     1921 tctagaggtg ctcttccagg ggcctgatcc cgatcccgag ggtgagtact aagcttcagc
     1981 gctcctgcct ggacgcatcc cggctatgca gccccagtcc agggcagcaa ggcaggcccc
     2041 gtctgcctct tcacccggag gcctctgccc gccccactca tgctcaggga gagggtcttc
     2101 tggctttttc cccaggctct gggcaggcac aggctaggtg cccctaaccc aggccctgca
     2161 cacaaagggg caggtgctgg gctcagacct gccaagagcc atatccggga ggaccctgcc
     2221 cctgacctaa gcccacccca aaggccaaac tctccactcc ctcagctcgg acaccttctc
     2281 tcctcccaga ttccagtaac tcccaatctt ctctctgcag agcccaaatc ttgtgacaaa
     2341 actcacacat gcccaccgtg cccaggtaag ccagcccagg cctcgccctc cagctcaagg
     2401 cgggacaggt gccctagagt agcctgcatc cagggacagg ccccagccgg gtgctgacac
     2461 gtccacctcc atctcttcct cagcacctga actcctgggg ggaccgtcag tcttcctctt
     2521 ccccccaaaa cccaaggaca ccctcatgat ctcccggacc cctgaggtca catgcgtggt
     2581 ggtggacgtg agccacgaag accctgaggt caagttcaac tggtacgtgg acggcgtgga
     2641 ggtgcataat gccaagacaa agccgcggga ggagcagtac aacagcacgt accgtgtggt
     2701 cagcgtcctc accgtcctgc accaggactg gctgaatggc aaggagtaca agtgcaaggt
     2761 ctccaacaaa gccctcccag cccccatcga gaaaaccatc tccaaagcca aaggtgggac
     2821 ccgtggggtg cgagggccac atggacagag gccggctcgg cccaccctct gccctgagag
     2881 tgaccgctgt accaacctct gtccctacag ggcagccccg agaaccacag gtgtacaccc
     2941 tgcccccatc ccgggatgag ctgaccaaga accaggtcag cctgacctgc ctggtcaaag
     3001 gcttctatcc cagcgacatc gccgtggagt gggagagcaa tgggcagccg gagaacaact
     3061 acaagaccac gcctcccgtg ctggactccg acggctcctt cttcctctac agcaagctca
     3121 ccgtggacaa gagcaggtgg cagcagggga acgtcttctc atgctccgtg atgcatgagg
     3181 ctctgcacaa ccactacacg cagaagagcc tctccctgtc tccgggtaaa tgagtgcgac
     3241 ggccgcgact ctagggtggc atccctgtga cccctcccca gtgcctctcc tggccctgga
     3301 agttgccact ccagtgccca ccagccttgt cctaataaaa ttaagttgca tcattttgtc
     3361 tgactaggtg tccttctata atattatggg gtggaggggg gtggtatgga gcaaggggca
     3421 agttgggaag acaacctgta gggcctgcgg ggtctattgg gaaccaagct ggagtgcagt
     3481 ggcacaatct tggctcactg caatctccgc ctcctgggtt caagcgattc tcctgcctca
     3541 gcctcccgag ttgttgggat tccaggcatg catgaccagg ctcagctaat ttttgttttt
     3601 ttggtagaga cggggtttca ccatattggc caggctggtc tccaactcct aatctcaggt
     3661 gatctaccca ccttggcctc ccaaattgct gggattacag gcgtgaacca ctgctccctt
     3721 ccctgtcctt ctgattttaa aataactata ccagcaggag gacgtccaga cacagcatag
     3781 gctacctggc catgcccaac cggtgggaca tttgagttgc ttgcttggca ctgtcctctc
     3841 atgcgttggg tccactcagt agatgcctgt tgaattgggt acgcggccag cttggctgtg
     3901 gaatgtgtgt cagttagggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca
     3961 aagcatgcat ctcaattagt cagcaaccag gtgtggaaag tccccaggct ccccagcagg
     4021 cagaagtatg caaagcatgc atctcaatta gtcagcaacc atagtcccgc ccctaactcc
     4081 gcccatcccg cccctaactc cgcccagttc cgcccattct ccgccccatg gctgactaat
     4141 tttttttatt tatgcagagg ccgaggccgc ctcggcctct gagctattcc agaagtagtg
     4201 aggaggcttt tttggaggcc taggcttttg caaaaagctc ctcgaggaac tgaaaaacca
     4261 gaaagttaat tccctatagt gagtcgtatt aaattcgtaa tcatgtcata gctgtttcct
     4321 gtgtgaaatt gttatccgct cacaattcca cacaacatac gagccggaag cataaagtgt
     4381 aaagcctggg gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc
     4441 gctttccagt cgggaaacct gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg
     4501 agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg
     4561 gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca
     4621 gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac
     4681 cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac
     4741 aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg
     4801 tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac
     4861 ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat
     4921 ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag
     4981 cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac
     5041 ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt
     5101 gctacatagt tcttgaagtg gtggcctaac tacggctaca ctagaagaac agtatttggt
     5161 atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc
     5221 aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga
     5281 aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac
     5341 gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc
     5401 cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct
     5461 gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca
     5521 tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct
     5581 ggccccagtg ctgcaatgat accgcgagac ccacgctcac cggctccaga tttatcagca
     5641 ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc
     5701 atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg
     5761 cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct
     5821 tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa
     5881 aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta
     5941 tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc
     6001 ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg
     6061 agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag aactttaaaa
     6121 gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg
     6181 agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc
     6241 accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg
     6301 gcgacacgga aatgttgaat actcatactc ttcctttttc aatattattg aagcatttat
     6361 cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata
     6421 ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg cgccctgtag cggcgcatta
     6481 agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag cgccctagcg
     6541 cccgctcctt tcgctttctt cccttccttt ctcgccacgt tcgccggctt tccccgtcaa
     6601 gctctaaatc gggggctccc tttagggttc cgatttagtg ctttacggca cctcgacccc
     6661 aaaaaacttg attagggtga tggttcacgt agtgggccat cgccctgata gacggttttt
     6721 cgccctttga cgttggagtc cacgttcttt aatagtggac tcttgttcca aactggaaca
     6781 acactcaacc ctatctcggt ctattctttt gatttataag ggattttgcc gatttcggcc
     6841 tattggttaa aaaatgagct gatttaacaa aaatttaacg cgaatttt
//
