LOCUS       sequence-260875-        6970 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of bioactive mouse IL-12 as a tandem dimer of p40 and p35
            linked with a Gly-Ser linker.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6970)
  AUTHORS   Nevil Singh
  TITLE     Nevil Singh IL-12 construct
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 6970)
  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: 
            "Unpublished"
COMMENT     Sequence Label: Tandem p40p35
FEATURES             Location/Qualifiers
     source          1..6970
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     intron          2802..3031
                     /label=chimeric intron
                     /note="chimera between introns from adenovirus and 
                     immunoglobulin heavy chain genes"
     primer_bind     3014..3033
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     misc_feature    join(3104..3678,3679..3681,3682..3690)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(3271..3288)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     3498..3517
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     regulatory      3706..3715
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(3712..3714,3715..3717,3718..4431)
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(3757..3778)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(4018..4037)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     4365..4386
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(4457..4474)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4463..4687
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     4734..4753
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4813..4829)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4813..4829)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4826..4848)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4837..4853
                     /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(4861..4867,4868..4885,4886..4891))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4906..4927
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5044..5061)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5215..5803)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5295..5314)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5974..6765,6766..6834))
                     /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     6597..6616
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6835..6939)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gcttggtacc
      901 gagctcggat cgatatctgc ggccgcgtcg ccaccatgtg tcctcagaag ctaaccatct
      961 cctggtttgc catcgttttg ctggtgtctc cactcatggc catgtgggag ctggagaaag
     1021 acgtttatgt tgtagaggtg gactggactc ccgatgcccc tggagaaaca gtgaacctca
     1081 cctgtgacac gcctgaagaa gatgacatca cctggacctc agaccagaga catggagtca
     1141 taggctctgg aaagaccctg accatcactg tcaaagagtt tctagatgct ggccagtaca
     1201 cctgccacaa aggaggcgag actctgagcc actcacatct gctgctccac aagaaggaaa
     1261 atggaatttg gtccactgaa attttaaaaa atttcaaaaa caagactttc ctgaagtgtg
     1321 aagcaccaaa ttactccgga cggttcacgt gctcatggct ggtgcaaaga aacatggact
     1381 tgaagttcaa catcaagagc agtagcagtt cccctgactc tcgggcagtg acatgtggaa
     1441 tggcgtctct gtctgcagag aaggtcacac tggaccaaag ggactatgag aagtattcag
     1501 tgtcctgcca ggaggatgtc acctgcccaa ctgccgagga gaccctgccc attgaactgg
     1561 cgttggaagc acggcagcag aataaatatg agaactacag caccagcttc ttcatcaggg
     1621 acatcatcaa accagacccg cccaagaact tgcagatgaa gcctttgaag aactcacagg
     1681 tggaggtcag ctgggagtac cctgactcct ggagcactcc ccattcctac ttctccctca
     1741 agttctttgt tcgaatccag cgcaagaaag aaaagatgaa ggagacagag gaggggtgta
     1801 accagaaagg tgcgttcctc gtagagaaga catctaccga agtccaatgc aaaggcggga
     1861 atgtctgcgt gcaagctcag gatcgctatt acaattcctc atgcagcaag tgggcatgtg
     1921 ttccctgcag ggtccgatcc ggcggcggcg ggagtggcgg cgggggttct ggcggaggcc
     1981 tcgctagcgg tggctccatg gtcagcgttc caacagcctc accctcggca tccagcagct
     2041 cctctcagtg ccggtccagc atgtgtcaat cacgctacct cctctttttg gccacccttg
     2101 ccctcctaaa ccacctcagt ttggccaggg tcattccagt ctctggacct gccaggtgtc
     2161 ttagccagtc ccgaaacctg ctgaagacca cagatgacat ggtgaagacg gccagagaaa
     2221 aactgaaaca ttattcctgc actgctgaag acatcgatca tgaagacatc acacgggacc
     2281 aaaccagcac attgaagacc tgtttaccac tggaactaca caagaacgag agttgcctgg
     2341 ctactagaga gacttcttcc acaacaagag ggagctgcct gcccccacag aagacgtctt
     2401 tgatgatgac cctgtgcctt ggtagcatct atgaggactt gaagatgtac cagacagagt
     2461 tccaggccat caacgcagca cttcagaatc acaaccatca gcagatcatt ctagacaagg
     2521 gcatgctggt ggccatcgat gagctgatgc agtctctgaa tcataatggc gagactctgc
     2581 gccagaaacc tcctgtggga gaagcagacc cttacagagt gaaaatgaag ctctgcatcc
     2641 tgcttcacgc cttcagcacc cgcgtcgtga ccatcaacag ggtgatgggc tatctgagct
     2701 ccgcctgagt taacgcgatc gcacggaatt cagtggatcc actagtaacg gccgccagtg
     2761 tgctggaatt aattcgctgt ctgcgagggc cagctgttgg ggtgagtact ccctctcaaa
     2821 agcgggcatg acttctgcgc taagattgtc agtttccaaa aacgaggagg atttgatatt
     2881 cacctggccc gcggtgatgc ctttgagggt ggccgcgtcc atctggtcag aaaagacaat
     2941 ctttttgttg tcaagcttga ggtgtggcag gcttgagatc tggccataca cttgagtgac
     3001 aatgacatcc actttgcctt tctctccaca ggtgtccact cccaggtcca actgcaggtc
     3061 gagcatgcat ctagggcggc caattccgcc cctctccccc ccccccctct ccctcccccc
     3121 cccctaacgt tactggccga agccgcttgg aataaggccg gtgtgcgttt gtctatatgt
     3181 tattttccac catattgccg tcttttggca atgtgagggc ccggaaacct ggccctgtct
     3241 tcttgacgag cattcctagg ggtctttccc ctctcgccaa aggaatgcaa ggtctgttga
     3301 atgtcgtgaa ggaagcagtt cctctggaag cttcttgaag acaaacaacg tctgtagcga
     3361 ccctttgcag gcagcggaac cccccacctg gcgacaggtg cctctgcggc caaaagccac
     3421 gtgtataaga tacacctgca aaggcggcac aaccccagtg ccacgttgtg agttggatag
     3481 ttgtggaaag agtcaaatgg ctctcctcaa gcgtattcaa caaggggctg aaggatgccc
     3541 agaaggtacc ccattgtatg ggatctgatc tggggcctcg gtgcacatgc tttacatgtg
     3601 tttagtcgag gttaaaaaaa cgtctaggcc ccccgaacca cggggacgtg gttttccttt
     3661 gaaaaacacg atgataagct tgccacaacc cgggatccac cggtcgccac catggtgagc
     3721 aagggcgagg agctgttcac cggggtggtg cccatcctgg tcgagctgga cggcgacgta
     3781 aacggccaca agttcagcgt gtccggcgag ggcgagggcg atgccaccta cggcaagctg
     3841 accctgaagt tcatctgcac caccggcaag ctgcccgtgc cctggcccac cctcgtgacc
     3901 accttcggct acggcctgca gtgcttcgcc cgctaccccg accacatgaa gcagcacgac
     3961 ttcttcaagt ccgccatgcc cgaaggctac gtccaggagc gcaccatctt cttcaaggac
     4021 gacggcaact acaagacccg cgccgaggtg aagttcgagg gcgacaccct ggtgaaccgc
     4081 atcgagctga agggcatcga cttcaaggag gacggcaaca tcctggggca caagctggag
     4141 tacaactaca acagccacaa cgtctatatc atggccgaca agcagaagaa cggcatcaag
     4201 gtgaacttca agatccgcca caacatcgag gacggcagcg tgcagctcgc cgaccactac
     4261 cagcagaaca cccccatcgg cgacggcccc gtgctgctgc ccgacaacca ctacctgagc
     4321 taccagtccg ccctgagcaa agaccccaac gagaagcgcg atcacatggt cctgctggag
     4381 ttcgtgaccg ccgccgggat cactctcggc atggacgagc tgtacaagta aagcggccct
     4441 agagctcgct gatcagcctc gactgtgcct tctagttgcc agccatctgt tgtttgcccc
     4501 tcccccgtgc cttccttgac cctggaaggt gccactccca ctgtcctttc ctaataaaat
     4561 gaggaaattg catcgcattg tctgagtagg tgtcattcta ttctgggggg tggggtgggg
     4621 caggacagca agggggagga ttgggaagac aatagcaggc atgctgggga tgcggtgggc
     4681 tctatggctt ctgaggcgga aagaaccagc tggggctcga gtgcattcta gttgtggttt
     4741 gtccaaactc atcaatgtat cttatcatgt ctgtataccg tcgacctcta gctagagctt
     4801 ggcgtaatca tggtcatagc tgtttcctgt gtgaaattgt tatccgctca caattccaca
     4861 caacatacga gccggaagca taaagtgtaa agcctggggt gcctaatgag tgagctaact
     4921 cacattaatt gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt cgtgccagct
     4981 gcattaatga atcggccaac gcgcggggag aggcggtttg cgtattgggc gctcttccgc
     5041 ttcctcgctc actgactcgc tgcgctcggt cgttcggctg cggcgagcgg tatcagctca
     5101 ctcaaaggcg gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg
     5161 agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca
     5221 taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa
     5281 cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc
     5341 tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc
     5401 gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct
     5461 gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg
     5521 tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag
     5581 gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta
     5641 cggctacact agaagaacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg
     5701 aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt
     5761 tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt
     5821 ttctacgggg tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag
     5881 attatcaaaa aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat
     5941 ctaaagtata tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc
     6001 tatctcagcg atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat
     6061 aactacgata cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc
     6121 acgctcaccg gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag
     6181 aagtggtcct gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag
     6241 agtaagtagt tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt
     6301 ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg
     6361 agttacatga tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt
     6421 tgtcagaagt aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc
     6481 tcttactgtc atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc
     6541 attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa
     6601 taccgcgcca catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg
     6661 aaaactctca aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc
     6721 caactgatct tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag
     6781 gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt
     6841 cctttttcaa tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt
     6901 tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc
     6961 acctgacgtc
//
