LOCUS       sequence-221702-        6477 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Liver-specific adeno-associated viral delivery and mammalian 
            expression of Flag-mRIPK1-S321E.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6477)
  AUTHORS   Geng J, Ito Y, Shi L, Amin P, Chu J, Ouchida AT, Mookhtiar AK, Zhao 
            H, Xu D, Shan B, Najafov A, Gao G, Akira S, Yuan J
  TITLE     Regulation of RIPK1 activation by TAK1-mediated phosphorylation 
            dictates apoptosis and necroptosis.
  JOURNAL   Nat Commun. 2017 Aug 25;8(1):359. doi: 10.1038/s41467-017-00406-w.
  PUBMED    28842570
REFERENCE   2  (bases 1 to 6477)
  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: "Nat 
            Commun."; date: "2017-08-25"; volume: "8(1):359. doi"; pages: " 
            10.1038/s41467-017-00406-w"
COMMENT     Sequence Label: pAAV2-TBG-FLAG-mRIPK1-S321E
FEATURES             Location/Qualifiers
     source          1..6477
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..410
                     /label=TBG promoter
                     /note="promoter from the human thyroxine-binding globulin 
                     gene (Yan et al., 2012)"
     intron          548..680
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     663..682
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     promoter        725..743
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             769..792
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     primer_bind     complement(2835..2854)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    2900..2955
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     repeat_region   3048..3177
                     /label=AAV2 ITR
     primer_bind     complement(3196..3213)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3196..3212)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3205..3227)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      3354..3809
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3441..3460)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3651..3672
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        3835..3939
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3940..4008,4009..4800)
                     /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(4158..4177)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4971..5559
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5460..5479
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     protein_bind    5847..5868
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(5883..5888,5889..5906,5907..5913)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5921..5937
                     /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     5926..5948
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     5945..5961
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5945..5961
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     repeat_region   5999..6128
                     /label=AAV2 ITR
ORIGIN
        1 tgcatgtata atttctacag aacctattag aaaggatcac ccagcctctg cttttgtaca
       61 actttccctt aaaaaactgc caattccact gctgtttggc ccaatagtga gaactttttc
      121 ctgctgcctc ttggtgcttt tgcctatggc ccctattctg cctgctgaag acactcttgc
      181 cagcatggac ttaaacccct ccagctctga caatcctctt tctcttttgt tttacatgaa
      241 gggtctggca gccaaagcaa tcactcaaag ttcaaacctt atcatttttt gctttgttcc
      301 tcttggcctt ggttttgtac atcagctttg aaaataccat cccagggtta atgctggggt
      361 taatttataa ctaagagtgc tctagttttg caatacagga catgctataa aaatggaaag
      421 atgttgcttt ctgagagaca gctttattgc ggtagtttat cacagttaaa ttgctaacgc
      481 agtcagtgct tctgacacaa cagtctcgaa cttaagctgc agaagttggt cgtgaggcac
      541 tgggcaggta agtatcaagg ttacaagaca ggtttaagga gaccaataga aactgggctt
      601 gtcgagacag agaagactct tgcgtttctg ataggcacct attggtctta ctgacatcca
      661 ctttgccttt ctctccacag gtgtccactc ccagttcaat tacagctctt aaggctagag
      721 tacttaatac gactcactat aggctagcct cgagaattca cgcgtatgga ctacaaagac
      781 gatgacgaca agggcatgca accagacatg tccttggaca atattaagat ggcatccagt
      841 gacctgctgg agaagacaga cctagacagc ggaggcttcg ggaaggtgtc cttgtgttac
      901 cacagaagcc atggatttgt catcctgaaa aaagtataca cagggcccaa ccgcgctgag
      961 tacaatgagg ttctcttgga agaggggaag atgatgcaca gactgagaca cagtcgagtg
     1021 gtgaagctac tgggcatcat catagaagaa gggaactatt cgctggtgat ggagtacatg
     1081 gagaagggca acctgatgca cgtgctaaag acccagatag atgtcccact ttcattgaaa
     1141 ggaaggataa tcgtggaggc catagaaggc atgtgctact tacatgacaa aggtgtgata
     1201 cacaaggacc tgaagcctga gaatatcctc gttgatcgtg actttcacat taagatagcc
     1261 gatcttggtg tggcttcctt taagacatgg agcaaactga ctaaggagaa agacaacaag
     1321 cagaaagaag tgagcagcac cactaagaag aacaatggtg gtacccttta ctacatggca
     1381 cccgaacacc tgaatgacat caatgcaaag cccacggaga agtcggacgt gtacagcttt
     1441 ggcattgtcc tttgggcaat atttgcaaaa aaggagccct atgagaatgt catctgtact
     1501 gagcagttcg tgatctgcat aaaatctggg aacaggccaa atgtagagga aatccttgag
     1561 tactgtccaa gggagatcat cagcctcatg gagcggtgct ggcaggcgat cccagaagac
     1621 aggccaacat ttcttggcat tgaagaagaa tttaggcctt tttacttaag tcattttgaa
     1681 gaatatgtag aagaggatgt ggcaagttta aagaaagagt atccagatca aagcccagtg
     1741 ctgcagagaa tgtttgaact ccagcatgac tgtgtaccct tacctccgag caggtcaaat
     1801 tcagaacaac ctggatcgct gcacagttcc caggggctcc agatgggtcc tgtggaggag
     1861 tcctggtttt cttcctcccc agagtaccca caggacgaga atgatcgcag tgtgcaggct
     1921 aagctgcaag aggaagccag ctatcatgct tttggaatat ttgcagagaa acagacaaaa
     1981 ccgcagccaa ggcagaatga ggcttacaac agagaggagg aaaggaaacg aagggtctct
     2041 catgacccct ttgcacagca gagagctcgt gagaatatta agagtgcagg agcaagaggt
     2101 cattctgatc ccagcacaac gagtcgtgga attgcagtgc aacagctgtc atggccagcc
     2161 acccaaacag tttggaacaa tggattgtat aatcagcatg gatttggaac tataggtaca
     2221 ggagtttggt atccgccaaa tctaagccaa atgtatagta cttataaaac tccagtgcca
     2281 gagaccaaca taccgggaag cacacccacc atgccatact tctctgggcc agtagcagat
     2341 gacctcataa aatatactat attcaatagt tctggtattc agattggaaa ccacaattat
     2401 atggatgttg gactgaattc acaaccacca aacaatactt gcaaagaaga gtcgacttcc
     2461 agacaccaag ccatctttga taacaccact agtctgactg atgaacacct gaaccctatc
     2521 agggaaaacc tgggaaggca gtggaaaaac tgtgcccgca agctgggctt cactgagtct
     2581 cagatcgatg aaatcgacca tgactatgaa agagatggac tgaaagagaa agtttaccaa
     2641 atgcttcaga agtggctgat gcgggaaggc accaaagggg ccacagtggg aaagttggcc
     2701 caggcacttc accaatgttg caggatagac ctgctgaacc acttgattcg tgccagccag
     2761 agctaaacgc gtggtacctc tagagtcgac ccgggcggcc tcgaggacgg ggtgaactac
     2821 gcctgaggat ccgatctttt tccctctgcc aaaaattatg gggacatcat gaagcccctt
     2881 gagcatctga cttctggcta ataaaggaaa tttattttca ttgcaatagt gtgttggaat
     2941 tttttgtgtc tctcactcgg aagcaattcg ttgatctgaa tttcgaccac ccataatacc
     3001 cattaccctg gtagataagt agcatggcgg gttaatcatt aactacaagg aacccctagt
     3061 gatggagttg gccactccct ctctgcgcgc tcgctcgctc actgaggccg ggcgaccaaa
     3121 ggtcgcccga cgcccgggct ttgcccgggc ggcctcagtg agcgagcgag cgcgcagcct
     3181 taattaacct aattcactgg ccgtcgtttt acaacgtcgt gactgggaaa accctggcgt
     3241 tacccaactt aatcgccttg cagcacatcc ccctttcgcc agctggcgta atagcgaaga
     3301 ggcccgcacc gatcgccctt cccaacagtt gcgcagcctg aatggcgaat gggacgcgcc
     3361 ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga ccgctacact
     3421 tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct tcctttctcg ccacgttcgc
     3481 cggctttccc cgtcaagctc taaatcgggg gctcccttta gggttccgat ttagtgcttt
     3541 acggcacctc gaccccaaaa aacttgatta gggtgatggt tcacgtagtg ggccatcgcc
     3601 ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg ttctttaata gtggactctt
     3661 gttccaaact ggaacaacac tcaaccctat ctcggtctat tcttttgatt tataagggat
     3721 tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa
     3781 ttttaacaaa atattaacgc ttacaattta ggtggcactt ttcggggaaa tgtgcgcgga
     3841 acccctattt gtttattttt ctaaatacat tcaaatatgt atccgctcat gagacaataa
     3901 ccctgataaa tgcttcaata atattgaaaa aggaagagta tgagtattca acatttccgt
     3961 gtcgccctta ttcccttttt tgcggcattt tgccttcctg tttttgctca cccagaaacg
     4021 ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac gagtgggtta catcgaactg
     4081 gatctcaaca gcggtaagat ccttgagagt tttcgccccg aagaacgttt tccaatgatg
     4141 agcactttta aagttctgct atgtggcgcg gtattatccc gtattgacgc cgggcaagag
     4201 caactcggtc gccgcataca ctattctcag aatgacttgg ttgagtactc accagtcaca
     4261 gaaaagcatc ttacggatgg catgacagta agagaattat gcagtgctgc cataaccatg
     4321 agtgataaca ctgcggccaa cttacttctg acaacgatcg gaggaccgaa ggagctaacc
     4381 gcttttttgc acaacatggg ggatcatgta actcgccttg atcgttggga accggagctg
     4441 aatgaagcca taccaaacga cgagcgtgac accacgatgc ctgtagcaat ggcaacaacg
     4501 ttgcgcaaac tattaactgg cgaactactt actctagctt cccggcaaca attaatagac
     4561 tggatggagg cggataaagt tgcaggacca cttctgcgct cggcccttcc ggctggctgg
     4621 tttattgctg ataaatctgg agccggtgag cgtgggtctc gcggtatcat tgcagcactg
     4681 gggccagatg gtaagccctc ccgtatcgta gttatctaca cgacggggag tcaggcaact
     4741 atggatgaac gaaatagaca gatcgctgag ataggtgcct cactgattaa gcattggtaa
     4801 ctgtcagacc aagtttactc atatatactt tagattgatt taaaacttca tttttaattt
     4861 aaaaggatct aggtgaagat cctttttgat aatctcatga ccaaaatccc ttaacgtgag
     4921 ttttcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc ttgagatcct
     4981 ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt
     5041 tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg
     5101 cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt caagaactct
     5161 gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc
     5221 gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg
     5281 tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa
     5341 ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg
     5401 gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg
     5461 ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga
     5521 tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt
     5581 ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc gttatcccct
     5641 gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg ccgcagccga
     5701 acgaccgagc gcagcgagtc agtgagcgag gaagcggaag agcgcccaat acgcaaaccg
     5761 cctctccccg cgcgttggcc gattcattaa tgcagctggc acgacaggtt tcccgactgg
     5821 aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc tcactcatta ggcaccccag
     5881 gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa ttgtgagcgg ataacaattt
     5941 cacacaggaa acagctatga ccatgattac gccagattta attaaggcct taattaggct
     6001 gcgcgctcgc tcgctcactg aggccgcccg ggcaaagccc gggcgtcggg cgacctttgg
     6061 tcgcccggcc tcagtgagcg agcgagcgcg cagagaggga gtggccaact ccatcactag
     6121 gggttccttg tagttaatga ttaacccgcc atgctactta tctaccaggg taatggggat
     6181 cctctagaac tatagctaga attcgccctt aagctagcag gttaattttt aaaaagcagt
     6241 caaaagtcca agtggccctt ggcagcattt actctctctg tttgctctgg ttaataatct
     6301 caggagcaca aacattccag atccaggtta atttttaaaa agcagtcaaa agtccaagtg
     6361 gcccttggca gcatttactc tctctgtttg ctctggttaa taatctcagg agcacaaaca
     6421 ttccagatcc ggcgcgccag ggctggaagc tacctttgac atcatttcct ctgcgaa
//
