LOCUS       sequence-214528-        6805 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of the indicated Flag- hRIPK1 mutant. Please 
            note Flag tag is N-terminal..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6805)
  TITLE     Yuan Lab Plasmids
REFERENCE   2  (bases 1 to 6805)
  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: pcDNA3.1-hRIPK1-Flag-delDD
FEATURES             Location/Qualifiers
     source          1..6805
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..380
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        381..584
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     535..555
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     629..648
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        629..647
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             691..714
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     promoter        complement(2505..2523)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(2506..2523)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(2543..2560)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2549..2773
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      complement(2905..3493)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2985..3004)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     polyA_signal    3822..3869
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     primer_bind     3894..3913
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     CDS             complement(4101..4895)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             complement(4101..4895)
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4212..4231)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     4822..4841
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     promoter        complement(4930..5287)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4944..5079
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     enhancer        5096..5287
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     5244..5264
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     CDS             complement(join(5347..6138,6139..6207))
                     /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     5970..5989
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6208..6312)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      6339..6794
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6426..6445)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6636..6657
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
ORIGIN
        1 gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
       61 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca
      121 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga
      181 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc
      241 aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
      301 ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat
      361 tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc
      421 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt
      481 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa
      541 tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctctctgg ctaactagag
      601 aacccactgc ttactggctt atcgaaatta atacgactca ctatagggag acccaagctt
      661 ggtaccggat cccccgggaa tcaaaacatg gattacaaag acgatgacga taaagaattc
      721 aggatgcaac cagacatgtc cttgaatgtc attaagatga aatccagtga cttcctggag
      781 agtgcagaac tggacagcgg aggctttggg aaggtgtctc tgtgtttcca cagaacccag
      841 ggactcatga tcatgaaaac agtgtacaag gggcccaact gcattgagca caacgaggcc
      901 ctcttggagg aggcgaagat gatgaacaga ctgagacaca gccgggtggt gaagctcctg
      961 ggcgtcatca tagaggaagg gaagtactcc ctggtgatgg agtacatgga gaagggcaac
     1021 ctgatgcacg tgctgaaagc cgagatgagt actccgcttt ctgtaaaagg aaggataatt
     1081 ttggaaatca ttgaaggaat gtgctactta catggaaaag gcgtgataca caaggacctg
     1141 aagcctgaaa atatccttgt tgataatgac ttccacatta agatcgcaga cctcggcctt
     1201 gcctccttta agatgtggag caaactgaat aatgaagagc acaatgagct gagggaagtg
     1261 gacggcaccg ctaagaagaa tggcggcacc ctctactaca tggcgcccga gcacctgaat
     1321 gacgtcaacg caaagcccac agagaagtcg gatgtgtaca gctttgctgt agtactctgg
     1381 gcgatatttg caaataagga gccatatgaa aatgctatct gtgagcagca gttgataatg
     1441 tgcataaaat ctgggaacag gccagatgtg gatgacatca ctgagtactg cccaagagaa
     1501 attatcagtc tcatgaagct ctgctgggaa gcgaatccgg aagctcggcc gacatttcct
     1561 ggcattgaag aaaaatttag gcctttttat ttaagtcaat tagaagaaag tgtagaagag
     1621 gacgtgaaga gtttaaagaa agagtattca aacgaaaatg cagttgtgaa gagaatgcag
     1681 tctcttcaac ttgattgtgt ggcagtacct tcaagccggt caaattcagc cacagaacag
     1741 cctggttcac tgcacagttc ccagggactt gggatgggtc ctgtggagga gtcctggttt
     1801 gctccttccc tggagcaccc acaagaagag aatgagccca gcctgcagag taaactccaa
     1861 gacgaagcca actaccatct ttatggcagc cgcatggaca ggcagacgaa acagcagccc
     1921 agacagaatg tggcttacaa cagagaggag gaaaggagac gcagggtctc ccatgaccct
     1981 tttgcacagc aaagacctta cgagaatttt cagaatacag agggaaaagg cactgcttat
     2041 tccagtgcag ccagtcatgg taatgcagtg caccagccct cagggctcac cagccaacct
     2101 caagtactgt atcagaacaa tggattatat agctcacatg gctttggaac aagaccactg
     2161 gatccaggaa cagcaggtcc cagagtttgg tacaggccaa ttccaagtca tatgcctagt
     2221 ctgcataata tcccagtgcc tgagaccaac tatctaggaa atacacccac catgccattc
     2281 agctccttgc caccaacaga tgaatctata aaatatacca tatacaatag tactggcatt
     2341 cagattggag cctacaatta tatggagatt ggtgggacga gttcatcact actagacagc
     2401 acaaatacga acttcaaaga agagccagct gctaagtacc aagctatctt tgataatacc
     2461 actagtctga cgctcgagta gatgactagt ctagagggcc ctattctata gtgtcaccta
     2521 aatgctagag ctcgctgatc agcctcgact gtgccttcta gttgccagcc atctgttgtt
     2581 tgcccctccc ccgtgccttc cttgaccctg gaaggtgcca ctcccactgt cctttcctaa
     2641 taaaatgagg aaattgcatc gcattgtctg agtaggtgtc attctattct ggggggtggg
     2701 gtggggcagg acagcaaggg ggaggattgg gaagacaata gcaggcatgc tggggatgcg
     2761 gtgggctcta tggcttctga ggcggaaaga accagtggcg gtaatacggt tatccacaga
     2821 atcaggggat aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg
     2881 taaaaaggcc gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa
     2941 aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt
     3001 tccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct
     3061 gtccgccttt ctcccttcgg gaagcgtggc gctttctcat agctcacgct gtaggtatct
     3121 cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc
     3181 cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt
     3241 atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc
     3301 tacagagttc ttgaagtggt ggcctaacta cggctacact agaagaacag tatttggtat
     3361 ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa
     3421 acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa
     3481 aaaaggatct caagaagatc ctttgatctt ttctacgggg tctgacgctc agtggaacga
     3541 aaactcacgt taagggattt tggtcatgag attatcaaaa aggatcttca cctagatcct
     3601 tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata tatgagtaac ctgaggctat
     3661 ggcagggcct gccgccccga cgttggctgc gagccctggg ccttcacccg aacttggggg
     3721 gtggggtggg gaaaaggaag aaacgcgggc gtattggccc caatggggtc tcggtggggt
     3781 atcgacagag tgccagccct gggaccgaac cccgcgttta tgaacaaacg acccaacacc
     3841 gtgcgtttta ttctgtcttt ttattgccgt catagcgcgg gttccttccg gtattgtctc
     3901 cttccgtgtt tcagttagcc tccccctagg gtgggcgaag aactccagca tgagatcccc
     3961 gcgctggagg atcatccagc cggcgtcccg gaaaacgatt ccgaagccca acctttcata
     4021 gaaggcggcg gtggaatcga aatctcgtga tggcaggttg ggcgtcgctt ggtcggtcat
     4081 ttcgaacccc agagtcccgc tcagaagaac tcgtcaagaa ggcgatagaa ggcgatgcgc
     4141 tgcgaatcgg gagcggcgat accgtaaagc acgaggaagc ggtcagccca ttcgccgcca
     4201 agctcttcag caatatcacg ggtagccaac gctatgtcct gatagcggtc cgccacaccc
     4261 agccggccac agtcgatgaa tccagaaaag cggccatttt ccaccatgat attcggcaag
     4321 caggcatcgc catgggtcac gacgagatcc tcgccgtcgg gcatgctcgc cttgagcctg
     4381 gcgaacagtt cggctggcgc gagcccctga tgctcttcgt ccagatcatc ctgatcgaca
     4441 agaccggctt ccatccgagt acgtgctcgc tcgatgcgat gtttcgcttg gtggtcgaat
     4501 gggcaggtag ccggatcaag cgtatgcagc cgccgcattg catcagccat gatggatact
     4561 ttctcggcag gagcaaggtg agatgacagg agatcctgcc ccggcacttc gcccaatagc
     4621 agccagtccc ttcccgcttc agtgacaacg tcgagcacag ctgcgcaagg aacgcccgtc
     4681 gtggccagcc acgatagccg cgctgcctcg tcttgcagtt cattcagggc accggacagg
     4741 tcggtcttga caaaaagaac cgggcgcccc tgcgctgaca gccggaacac ggcggcatca
     4801 gagcagccga ttgtctgttg tgcccagtca tagccgaata gcctctccac ccaagcggcc
     4861 ggagaacctg cgtgcaatcc atcttgttca atcatgcgaa acgatcctca tcctgtctct
     4921 tgatcgatct ttgcaaaagc ctaggcctcc aaaaaagcct cctcactact tctggaatag
     4981 ctcagaggcc gaggcggcct cggcctctgc ataaataaaa aaaattagtc agccatgggg
     5041 cggagaatgg gcggaactgg gcggagttag gggcgggatg ggcggagtta ggggcgggac
     5101 tatggttgct gactaattga gatgcatgct ttgcatactt ctgcctgctg gggagcctgg
     5161 ggactttcca cacctggttg ctgactaatt gagatgcatg ctttgcatac ttctgcctgc
     5221 tggggagcct ggggactttc cacaccctaa ctgacacaca ttccacagct ggttctttcc
     5281 gcctcaggac tcttcctttt tcaataaatc aatctaaagt atatatgagt aaacttggtc
     5341 tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc tatttcgttc
     5401 atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg gcttaccatc
     5461 tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag atttatcagc
     5521 aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt tatccgcctc
     5581 catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag ttaatagttt
     5641 gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc
     5701 ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca tgttgtgcaa
     5761 aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt
     5821 atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat ccgtaagatg
     5881 cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta tgcggcgacc
     5941 gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca gaactttaaa
     6001 agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct taccgctgtt
     6061 gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat cttttacttt
     6121 caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa agggaataag
     6181 ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt gaagcattta
     6241 tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa ataaacaaat
     6301 aggggttccg cgcacatttc cccgaaaagt gccacctgac gcgccctgta gcggcgcatt
     6361 aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca gcgccctagc
     6421 gcccgctcct ttcgctttct tcccttcctt tctcgccacg ttcgccggct ttccccgtca
     6481 agctctaaat cgggggctcc ctttagggtt ccgatttagt gctttacggc acctcgaccc
     6541 caaaaaactt gattagggtg atggttcacg tagtgggcca tcgccctgat agacggtttt
     6601 tcgccctttg acgttggagt ccacgttctt taatagtgga ctcttgttcc aaactggaac
     6661 aacactcaac cctatctcgg tctattcttt tgatttataa gggattttgc cgatttcggc
     6721 ctattggtta aaaaatgagc tgatttaaca aaaatttaac gcgaatttta acaaaatatt
     6781 aacgcttaca atttacgcgc gcgtt
//
