LOCUS       sequence-252081-        8197 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses human alpha-2M (from S24 to N1473) in insect cells. With 
            C-t H6x tag..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8197)
  AUTHORS   Marino-Puertas L, Del Amo-Maestro L, Taules M, Gomis-Ruth FX, Goulas
            T
  TITLE     Recombinant production of human alpha2-macroglobulin variants and 
            interaction studies with recombinant G-related alpha2-macroglobulin 
            binding protein and latent transforming growth factor-beta2.
  JOURNAL   Sci Rep. 2019 Jun 24;9(1):9186. doi: 10.1038/s41598-019-45712-z.
  PUBMED    31235767
REFERENCE   2  (bases 1 to 8197)
  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: "Sci Rep."; 
            date: "2019-06-24"; volume: "9(1):9186. doi"; pages: " 
            10.1038/s41598-019-45712-z"
COMMENT     Sequence Label: pIE-h-alpha-2M-H6
FEATURES             Location/Qualifiers
     source          1..8197
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..592
                     /label=IE1 promoter
                     /note="promoter of the ie1 gene from the baculovirus 
                     Autographa californica"
     CDS             5014..5031
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             5074..5091
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             5120..5164
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     CDS             5186..5209
                     /codon_start=1
                     /product="8xHis affinity tag"
                     /label=8xHis
                     /translation="HHHHHHHH"
     terminator      5228..5535
                     /label=IE1 terminator
                     /note="terminator of the ie1 gene from the baculovirus 
                     Autographa californica"
     promoter        5565..5669
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5670..5738,5739..6530)
                     /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     complement(5888..5907)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6701..7289
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7190..7209
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7443..7460
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    7577..7598
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(7613..7618,7619..7636,7637..7643)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7651..7667
                     /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     7656..7678
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     7675..7691
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     7675..7691
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     enhancer        7712..8194
                     /gene="Autographa californica hr5"
                     /label=hr5 enhancer
                     /note="baculovirus early transcription enhancer"
ORIGIN
        1 cgatgtcttt gtgatgcgcg cgacattttt gtaggttatt gataaaatga acggatacgt
       61 tgcccgacat tatcattaaa tccttggcgt agaatttgtc gggtccattg tccgtgtgcg
      121 ctagcatgcc cgtaacggac ctcgtacttt tggcttcaaa ggttttgcgc acagacaaaa
      181 tgtgccacac ttgcagctct gcatgtgtgc gcgttaccac aaatcccaac ggcgcagtgt
      241 acttgttgta tgcaaataaa tctcgataaa ggcgcggcgc gcgaatgcag ctgatcacgt
      301 acgctcctcg tgttccgttc aaggacggtg ttatcgacct cagattaatg tttatcggcc
      361 gactgttttc gtatccgctc accaaacgcg tttttgcatt aacattgtat gtcggcggat
      421 gttctatatc taatttgaat aaataaacga taaccgcgtt ggttttagag ggcataataa
      481 aagaaatatt gttatcgtgt tcgccattag ggcagtataa attgacgttc atgttggata
      541 ttgtttcagt tgcaagttga cactggcggc gacaagatcg tgaacaacca agtgactatg
      601 tacaagctca cagtcttcct gatgttcatc gctttcgtca tcatcgctga ggcctcagtc
      661 tctggaaaac cgcagtatat ggttctggtc ccctccctgc tccacactga gaccactgag
      721 aagggctgtg tccttctgag ctacctgaat gagacagtga ctgtaagtgc ttccttggag
      781 tctgtcaggg gaaacaggag cctcttcact gacctggagg cggagaatga cgtactccac
      841 tgtgtcgcct tcgctgtccc aaagtcttca tccaatgagg aggtaatgtt cctcactgtc
      901 caagtgaaag gaccaaccca agaatttaag aagcggacca cagtgatggt taagaacgag
      961 gacagtctgg tctttgtcca gacagacaaa tcaatctaca aaccagggca gacagtgaaa
     1021 tttcgtgttg tctccatgga tgaaaacttt caccccctga atgagttgat tccactagta
     1081 tacattcagg atcccaaagg aaatcgcatc gcacaatggc agagtttcca gttagagggt
     1141 ggcctcaagc aattttcttt tcccctctca tcagagccct tccagggctc ctacaaggtg
     1201 gtggtacaga agaaatcagg tggaaggaca gagcaccctt tcaccgtgga ggaatttgtt
     1261 cttcccaagt ttgaagtaca agtaacagtg ccaaagataa tcaccatctt ggaagaagag
     1321 atgaatgtat cagtgtgtgg cctatacaca tatgggaagc ctgtccctgg acatgtgact
     1381 gtgagcattt gcagaaagta tagtgacgct tccgactgcc acggtgaaga ttcacaggct
     1441 ttctgtgaga aattcagtgg acagctaaac agccatggct gcttctatca gcaagtaaaa
     1501 accaaggtct tccagctgaa gaggaaggag tatgaaatga aacttcacac tgaggcccag
     1561 atccaagaag aaggaacagt ggtggaattg actggaaggc agtccagtga aatcacaaga
     1621 accataacca aactctcatt tgtgaaagtg gactcacact ttcgacaggg aattcccttc
     1681 tttgggcagg tgcgcctagt agatgggaaa ggcgtcccta taccaaataa agtcatattc
     1741 atcagaggaa atgaagcaaa ctattactcc aatgctacca cggatgagca tggccttgta
     1801 cagttctcta tcaacaccac caatgttatg ggtacctctc ttactgttag ggtcaattac
     1861 aaggatcgta gtccctgtta cggctaccag tgggtgtcag aagaacacga agaggcacat
     1921 cacactgctt atcttgtgtt ctccccaagc aagagctttg tccaccttga gcccatgtct
     1981 catgaactac cctgtggcca tactcagaca gtccaggcac attatattct gaatggaggc
     2041 accctgctgg ggctgaagaa gctctccttc tattatctga taatggcaaa gggaggcatt
     2101 gtccgaactg ggactcatgg actgcttgtg aagcaggaag acatgaaggg ccatttttcc
     2161 atctcaatcc ctgtgaagtc agacattgct cctgtcgctc ggttgctcat ctatgctgtt
     2221 ttacctaccg gggacgtgat tggggattct gcaaaatatg atgttgaaaa ttgtctggcc
     2281 aacaaggtgg atttgagctt cagcccatca caaagtctcc cagcctcaca cgcccacctg
     2341 cgagtcacag cggctcctca gtccgtctgc gccctccgtg ctgtggacca aagcgtgctg
     2401 ctcatgaagc ctgatgctga gctctcggcg tcctcggttt acaacctgct accagaaaag
     2461 gacctcactg gcttccctgg gcctttgaat gaccaggacg atgaagactg catcaatcgt
     2521 cataatgtct atattaatgg aatcacatat actccagtat caagtacaaa tgaaaaggat
     2581 atgtacagct tcctagagga catgggctta aaggcattca ccaactcaaa gattcgtaaa
     2641 cccaaaatgt gtccacagct tcaacagtat gaaatgcatg gacctgaagg tctacgtgta
     2701 ggtttttatg agtcagatgt aatgggaaga ggccatgcac gcctggtgca tgttgaagag
     2761 cctcacacgg agaccgtacg aaagtacttc cctgagacat ggatctggga tttggtggtg
     2821 gtaaactcag caggtgtggc tgaggtagga gtaacagtcc ctgacaccat caccgagtgg
     2881 aaggcagggg ccttctgcct gtctgaagat gctggacttg gtatctcttc cactgcctct
     2941 ctccgagcct tccagccctt ctttgtggag ctcacaatgc cttactctgt gattcgtgga
     3001 gaggccttca cactcaaggc cacggtccta aactaccttc ccaaatgcat ccgggtcagt
     3061 gtgcagctgg aagcctctcc cgccttccta gctgtcccag tggagaagga acaagcgcct
     3121 cactgcatct gtgcaaacgg gcggcaaact gtgtcctggg cagtaacccc aaagtcatta
     3181 ggaaatgtga atttcactgt gagcgcagag gcactagagt ctcaagagct gtgtgggact
     3241 gaggtgcctt cagttcctga acacggaagg aaagacacag tcatcaagcc tctgttggtt
     3301 gaacctgaag gactagagaa ggaaacaaca ttcaactccc tactttgtcc atcaggtggt
     3361 gaggtttctg aagaattatc cctgaaactg ccaccaaatg tggtagaaga atctgcccga
     3421 gcttctgtct cagttttggg agacatatta ggctctgcca tgcaaaacac acaaaatctt
     3481 ctccagatgc cctatggctg tggagagcag aatatggtcc tctttgctcc taacatctat
     3541 gtactggatt atctaaatga aacacagcag cttactccag aggtcaagtc caaggccatt
     3601 ggctatctca acactggtta ccagagacag ttgaactaca aacactatga tggctcctac
     3661 agcacctttg gggagcgata tggcaggaac cagggcaaca cctggctcac agcctttgtt
     3721 ctgaagactt ttgcccaagc tcgagcctac atcttcatcg atgaagcaca cattacccaa
     3781 gccctcatat ggctctccca gaggcagaag gacaatggct gtttcaggag ctctgggtca
     3841 ctgctcaaca atgccataaa gggaggagta gaagatgaag tgaccctctc cgcctatatc
     3901 accatcgccc ttctggagat tcctctcaca gtcactcacc ctgttgtccg caatgccctg
     3961 ttttgcctgg agtcagcctg gaagacagca caagaagggg accatggcag ccatgtatat
     4021 accaaagcac tgctggccta tgcttttgcc ctggcaggta accaggacaa gaggaaggaa
     4081 gtactcaagt cacttaatga ggaagctgtg aagaaagaca actctgtcca ttgggagcgc
     4141 cctcagaaac ccaaggcacc agtggggcat ttttacgaac cccaggctcc ctctgctgag
     4201 gtggagatga catcctatgt gctcctcgct tatctcacgg cccagccagc cccaacctcg
     4261 gaggacctga cctctgcaac caacatcgtg aagtggatca cgaagcagca gaatgcccag
     4321 ggcggtttct cctccaccca ggacacagtg gtggctctcc atgctctgtc caaatatgga
     4381 gcagccacat ttaccaggac tgggaaggct gcacaggtga ctatccagtc ttcagggaca
     4441 ttttccagca aattccaagt ggacaacaac aaccgcctgt tactgcagca ggtctcattg
     4501 ccagagctgc ctggggaata cagcatgaaa gtgacaggag aaggatgtgt ctacctccag
     4561 acatccttga aatacaatat tctcccagaa aaggaagagt tcccctttgc tttaggagtg
     4621 cagactctgc ctcaaacttg tgatgaaccc aaagcccaca ccagcttcca aatctcccta
     4681 agtgtcagtt acacagggag ccgctctgcc tccaacatgg cgatcgttga tgtgaagatg
     4741 gtctctggct tcattcccct gaagccaaca gtgaaaatgc ttgaaagatc taaccatgtg
     4801 agccggacag aagtcagcag caaccatgtc ttgatttacc ttgataaggt gtcaaatcag
     4861 acactgagct tgttcttcac ggttctgcaa gatgtcccag taagagatct gaaaccagcc
     4921 atagtgaaag tctatgatta ctacgagacg gatgagtttg caattgctga gtacaatgct
     4981 ccttgcagca aagatcttgg aaatcctacc ggtcatcatc accatcacca ttgagtttgg
     5041 caaggccgga tccgttctac cctgcgtcac gtgcatcacc atcaccatca ctaataagct
     5101 tgcggccgca tcttctggta aagaaaccgc tgctgcgaaa tttgaacgcc agcacatgga
     5161 ctcgccaccg ccttctggcc tcgagcacca ccatcaccat caccatcact aagtgattaa
     5221 cctcaggtta tacatatatt ttgaatttaa ttaattatac atatatttta tattattttt
     5281 gtcttttatt atcgaggggc cgttgttggt gtggggtttt gcatagaaat aacaatggga
     5341 gttggcgacg ttgctgcgcc aacaccacct cccttccctc ctttcatcat gtatctgtag
     5401 ataaaataaa atattaaacc taaaaacaag accgcgccta tcaacaaaat gataggcatt
     5461 aacttgccgc tgacgctgtc actaacgttg gacgatttgc cgactaaacc ttcatcgccc
     5521 agtaaccaat ctagacgtca ggtggcactt ttcggggaaa tgtgcgcgga acccctattt
     5581 gtttattttt ctaaatacat tcaaatatgt atccgctcat gagacaataa ccctgataaa
     5641 tgcttcaata atattgaaaa aggaagagta tgagtattca acatttccgt gtcgccctta
     5701 ttcccttttt tgcggcattt tgccttcctg tttttgctca cccagaaacg ctggtgaaag
     5761 taaaagatgc tgaagatcag ttgggtgcac gagtgggtta catcgaactg gatctcaaca
     5821 gcggtaagat ccttgagagt tttcgccccg aagaacgttt tccaatgatg agcactttta
     5881 aagttctgct atgtggcgcg gtattatccc gtattgacgc cgggcaagag caactcggtc
     5941 gccgcataca ctattctcag aatgacttgg ttgagtactc accagtcaca gaaaagcatc
     6001 ttacggatgg catgacagta agagaattat gcagtgctgc cataaccatg agtgataaca
     6061 ctgcggccaa cttacttctg acaacgatcg gaggaccgaa ggagctaacc gcttttttgc
     6121 acaacatggg ggatcatgta actcgccttg atcgttggga accggagctg aatgaagcca
     6181 taccaaacga cgagcgtgac accacgatgc ctgtagcaat ggcaacaacg ttgcgcaaac
     6241 tattaactgg cgaactactt actctagctt cccggcaaca attaatagac tggatggagg
     6301 cggataaagt tgcaggacca cttctgcgct cggcccttcc ggctggctgg tttattgctg
     6361 ataaatctgg agccggtgag cgtgggtctc gcggtatcat tgcagcactg gggccagatg
     6421 gtaagccctc ccgtatcgta gttatctaca cgacggggag tcaggcaact atggatgaac
     6481 gaaatagaca gatcgctgag ataggtgcct cactgattaa gcattggtaa ctgtcagacc
     6541 aagtttactc atatatactt tagattgatt taaaacttca tttttaattt aaaaggatct
     6601 aggtgaagat cctttttgat aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc
     6661 actgagcgtc agaccccgta gaaaagatca aaggatcttc ttgagatcct ttttttctgc
     6721 gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt tgtttgccgg
     6781 atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg cagataccaa
     6841 atactgttct tctagtgtag ccgtagttag gccaccactt caagaactct gtagcaccgc
     6901 ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc gataagtcgt
     6961 gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg tcgggctgaa
     7021 cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa ctgagatacc
     7081 tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc
     7141 cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg ggaaacgcct
     7201 ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga tttttgtgat
     7261 gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt ttacggttcc
     7321 tggccttttg ctggcctttt gctcacatgt tctttcctgc gttatcccct gattctgtgg
     7381 ataaccgtat taccgccttt gagtgagctg ataccgctcg ccgcagccga acgaccgagc
     7441 gcagcgagtc agtgagcgag gaagcggaag agcgcccaat acgcaaaccg cctctccccg
     7501 cgcgttggcc gattcattaa tgcagctggc acgacaggtt tcccgactgg aaagcgggca
     7561 gtgagcgcaa cgcaattaat gtgagttagc tcactcatta ggcaccccag gctttacact
     7621 ttatgcttcc ggctcgtatg ttgtgtggaa ttgtgagcgg ataacaattt cacacaggaa
     7681 acagctatga ccatgattac gaattcccgg gcgcgtaaaa cacaatcaag tatgagtcat
     7741 aagctgatgt catgttttgc acacggctca taaccgaact ggctttacga gtagaattct
     7801 acttgtaacg cacgatcagt ggatgatgtc atttgttttt caaatcgaga tgatgtcatg
     7861 ttttgcacac ggctcataaa ctcgctttac gagtagaatt ctacgtgtaa cgcacgatcg
     7921 attgatgagt catttgtttt gcaatatgat atcatacaat atgactcatt tgtttttcaa
     7981 aaccgaactt gatttacggg tagaattcta cttgtaaagc acaatcaaaa agatgatgtc
     8041 atttgttttt caaaactgaa ctcgctttac gagtagaatt ctacgtgtaa aacacaatca
     8101 agaaatgatg tcatttgtta taaaaataaa agctgatgtc atgttttgca catggctcat
     8161 aactaaactc gctttacggg tagaattcta cgcgcgt
//
