LOCUS       sequence-252071-        5908 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Expresses human C-alpha-2M (from R787 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 5908)
  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 5908)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 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-C-h-alpha-2M-H6
FEATURES             Location/Qualifiers
     source          1..5908
                     /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             2725..2742
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             2785..2802
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             2831..2875
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     CDS             2897..2920
                     /codon_start=1
                     /product="8xHis affinity tag"
                     /label=8xHis
                     /translation="HHHHHHHH"
     terminator      2939..3246
                     /label=IE1 terminator
                     /note="terminator of the ie1 gene from the baculovirus 
                     Autographa californica"
     promoter        3276..3380
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3381..3449,3450..4241)
                     /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(3599..3618)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4412..5000
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4901..4920
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5154..5171
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    5288..5309
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(5324..5329,5330..5347,5348..5354)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5362..5378
                     /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     5367..5389
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     5386..5402
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5386..5402
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     enhancer        5423..5905
                     /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 ggcccgagcc
      661 ttccagccct tctttgtgga gctcacaatg ccttactctg tgattcgtgg agaggccttc
      721 acactcaagg ccacggtcct aaactacctt cccaaatgca tccgggtcag tgtgcagctg
      781 gaagcctctc ccgccttcct agctgtccca gtggagaagg aacaagcgcc tcactgcatc
      841 tgtgcaaacg ggcggcaaac tgtgtcctgg gcagtaaccc caaagtcatt aggaaatgtg
      901 aatttcactg tgagcgcaga ggcactagag tctcaagagc tgtgtgggac tgaggtgcct
      961 tcagttcctg aacacggaag gaaagacaca gtcatcaagc ctctgttggt tgaacctgaa
     1021 ggactagaga aggaaacaac attcaactcc ctactttgtc catcaggtgg tgaggtttct
     1081 gaagaattat ccctgaaact gccaccaaat gtggtagaag aatctgcccg agcttctgtc
     1141 tcagttttgg gagacatatt aggctctgcc atgcaaaaca cacaaaatct tctccagatg
     1201 ccctatggct gtggagagca gaatatggtc ctctttgctc ctaacatcta tgtactggat
     1261 tatctaaatg aaacacagca gcttactcca gaggtcaagt ccaaggccat tggctatctc
     1321 aacactggtt accagagaca gttgaactac aaacactatg atggctccta cagcaccttt
     1381 ggggagcgat atggcaggaa ccagggcaac acctggctca cagcctttgt tctgaagact
     1441 tttgcccaag ctcgagccta catcttcatc gatgaagcac acattaccca agccctcata
     1501 tggctctccc agaggcagaa ggacaatggc tgtttcagga gctctgggtc actgctcaac
     1561 aatgccataa agggaggagt agaagatgaa gtgaccctct ccgcctatat caccatcgcc
     1621 cttctggaga ttcctctcac agtcactcac cctgttgtcc gcaatgccct gttttgcctg
     1681 gagtcagcct ggaagacagc acaagaaggg gaccatggca gccatgtata taccaaagca
     1741 ctgctggcct atgcttttgc cctggcaggt aaccaggaca agaggaagga agtactcaag
     1801 tcacttaatg aggaagctgt gaagaaagac aactctgtcc attgggagcg ccctcagaaa
     1861 cccaaggcac cagtggggca tttttacgaa ccccaggctc cctctgctga ggtggagatg
     1921 acatcctatg tgctcctcgc ttatctcacg gcccagccag ccccaacctc ggaggacctg
     1981 acctctgcaa ccaacatcgt gaagtggatc acgaagcagc agaatgccca gggcggtttc
     2041 tcctccaccc aggacacagt ggtggctctc catgctctgt ccaaatatgg agcagccaca
     2101 tttaccagga ctgggaaggc tgcacaggtg actatccagt cttcagggac attttccagc
     2161 aaattccaag tggacaacaa caaccgcctg ttactgcagc aggtctcatt gccagagctg
     2221 cctggggaat acagcatgaa agtgacagga gaaggatgtg tctacctcca gacatccttg
     2281 aaatacaata ttctcccaga aaaggaagag ttcccctttg ctttaggagt gcagactctg
     2341 cctcaaactt gtgatgaacc caaagcccac accagcttcc aaatctccct aagtgtcagt
     2401 tacacaggga gccgctctgc ctccaacatg gcgatcgttg atgtgaagat ggtctctggc
     2461 ttcattcccc tgaagccaac agtgaaaatg cttgaaagat ctaaccatgt gagccggaca
     2521 gaagtcagca gcaaccatgt cttgatttac cttgataagg tgtcaaatca gacactgagc
     2581 ttgttcttca cggttctgca agatgtccca gtaagagatc tgaaaccagc catagtgaaa
     2641 gtctatgatt actacgagac ggatgagttt gcaattgctg agtacaatgc tccttgcagc
     2701 aaagatcttg gaaatcctac cggtcatcat caccatcacc attgagtttg gcaaggccgg
     2761 atccgttcta ccctgcgtca cgtgcatcac catcaccatc actaataagc ttgcggccgc
     2821 atcttctggt aaagaaaccg ctgctgcgaa atttgaacgc cagcacatgg actcgccacc
     2881 gccttctggc ctcgagcacc accatcacca tcaccatcac taagtgatta acctcaggtt
     2941 atacatatat tttgaattta attaattata catatatttt atattatttt tgtcttttat
     3001 tatcgagggg ccgttgttgg tgtggggttt tgcatagaaa taacaatggg agttggcgac
     3061 gttgctgcgc caacaccacc tcccttccct cctttcatca tgtatctgta gataaaataa
     3121 aatattaaac ctaaaaacaa gaccgcgcct atcaacaaaa tgataggcat taacttgccg
     3181 ctgacgctgt cactaacgtt ggacgatttg ccgactaaac cttcatcgcc cagtaaccaa
     3241 tctagacgtc aggtggcact tttcggggaa atgtgcgcgg aacccctatt tgtttatttt
     3301 tctaaataca ttcaaatatg tatccgctca tgagacaata accctgataa atgcttcaat
     3361 aatattgaaa aaggaagagt atgagtattc aacatttccg tgtcgccctt attccctttt
     3421 ttgcggcatt ttgccttcct gtttttgctc acccagaaac gctggtgaaa gtaaaagatg
     3481 ctgaagatca gttgggtgca cgagtgggtt acatcgaact ggatctcaac agcggtaaga
     3541 tccttgagag ttttcgcccc gaagaacgtt ttccaatgat gagcactttt aaagttctgc
     3601 tatgtggcgc ggtattatcc cgtattgacg ccgggcaaga gcaactcggt cgccgcatac
     3661 actattctca gaatgacttg gttgagtact caccagtcac agaaaagcat cttacggatg
     3721 gcatgacagt aagagaatta tgcagtgctg ccataaccat gagtgataac actgcggcca
     3781 acttacttct gacaacgatc ggaggaccga aggagctaac cgcttttttg cacaacatgg
     3841 gggatcatgt aactcgcctt gatcgttggg aaccggagct gaatgaagcc ataccaaacg
     3901 acgagcgtga caccacgatg cctgtagcaa tggcaacaac gttgcgcaaa ctattaactg
     3961 gcgaactact tactctagct tcccggcaac aattaataga ctggatggag gcggataaag
     4021 ttgcaggacc acttctgcgc tcggcccttc cggctggctg gtttattgct gataaatctg
     4081 gagccggtga gcgtgggtct cgcggtatca ttgcagcact ggggccagat ggtaagccct
     4141 cccgtatcgt agttatctac acgacgggga gtcaggcaac tatggatgaa cgaaatagac
     4201 agatcgctga gataggtgcc tcactgatta agcattggta actgtcagac caagtttact
     4261 catatatact ttagattgat ttaaaacttc atttttaatt taaaaggatc taggtgaaga
     4321 tcctttttga taatctcatg accaaaatcc cttaacgtga gttttcgttc cactgagcgt
     4381 cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct
     4441 gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc
     4501 taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgttc
     4561 ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc
     4621 tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg
     4681 ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt
     4741 cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg
     4801 agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg
     4861 gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc tggtatcttt
     4921 atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag
     4981 gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc ctggcctttt
     5041 gctggccttt tgctcacatg ttctttcctg cgttatcccc tgattctgtg gataaccgta
     5101 ttaccgcctt tgagtgagct gataccgctc gccgcagccg aacgaccgag cgcagcgagt
     5161 cagtgagcga ggaagcggaa gagcgcccaa tacgcaaacc gcctctcccc gcgcgttggc
     5221 cgattcatta atgcagctgg cacgacaggt ttcccgactg gaaagcgggc agtgagcgca
     5281 acgcaattaa tgtgagttag ctcactcatt aggcacccca ggctttacac tttatgcttc
     5341 cggctcgtat gttgtgtgga attgtgagcg gataacaatt tcacacagga aacagctatg
     5401 accatgatta cgaattcccg ggcgcgtaaa acacaatcaa gtatgagtca taagctgatg
     5461 tcatgttttg cacacggctc ataaccgaac tggctttacg agtagaattc tacttgtaac
     5521 gcacgatcag tggatgatgt catttgtttt tcaaatcgag atgatgtcat gttttgcaca
     5581 cggctcataa actcgcttta cgagtagaat tctacgtgta acgcacgatc gattgatgag
     5641 tcatttgttt tgcaatatga tatcatacaa tatgactcat ttgtttttca aaaccgaact
     5701 tgatttacgg gtagaattct acttgtaaag cacaatcaaa aagatgatgt catttgtttt
     5761 tcaaaactga actcgcttta cgagtagaat tctacgtgta aaacacaatc aagaaatgat
     5821 gtcatttgtt ataaaaataa aagctgatgt catgttttgc acatggctca taactaaact
     5881 cgctttacgg gtagaattct acgcgcgt
//
