LOCUS       sequence-340120-        6358 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses HA-tagged CDK2 in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6358)
  AUTHORS   Simoneschi D, Rona G, Zhou N, Jeong YT, Jiang S, Milletti G, Arbini 
            AA, O'Sullivan A, Wang AA, Nithikasem S, Keegan S, Siu Y, 
            Cianfanelli V, Maiani E, Nazio F, Cecconi F, Boccalatte F, Fenyo D, 
            Jones DR, Busino L, Pagano M
  TITLE     CRL4(AMBRA1) is a master regulator of D-type cyclins.
  JOURNAL   Nature. 2021 Apr;592(7856):789-793. doi: 10.1038/s41586-021-03445-y.
            Epub 2021 Apr 14.
  PUBMED    33854235
REFERENCE   2  (bases 1 to 6358)
  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"; doi: 
            "10.1038/s41586-021-03445-y"; journalName: "Nature"; date: 
            "2021-04"; volume: "592"; issue: "7856"; pages: "789-793"
COMMENT     Sequence Label: pcDNA3.1-HA-CDK2
FEATURES             Location/Qualifiers
     source          1..6358
                     /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"
     CDS             904..930
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /note="This is a variant of the standard HA tag, which has 
                     sequence YPYDVPDYA. Some anti-HA antibodies may not 
                     recognize this variant."
                     /translation="YPYDVPDYA"
     CDS             904..930
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             910..936
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /note="This is a variant of the standard HA tag, which has 
                     sequence YPYDVPDYA. Some anti-HA antibodies may not 
                     recognize this variant."
                     /translation="YDVPDYASL"
     promoter        complement(1911..1929)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(1912..1929)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(1949..1966)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1955..2179
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2225..2653
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2312..2331)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2522..2543
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2662..2682)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2667..2996
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2847..2982
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2909..2928
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3063..3857
                     /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
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3117..3136)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3727..3746
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4031..4152
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4068..4087)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4122..4141
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4201..4217)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4201..4217)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4214..4236)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4225..4241
                     /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(4249..4255,4256..4273,4274..4279))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4294..4315
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4432..4449)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4603..5191)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4683..4702)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5362..6153,6154..6222))
                     /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     5985..6004
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6223..6327)
                     /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 gcttaccatg
      901 gcctacccct acgacgtgcc cgactacgcc tccctcggat ccagtgtggt ggaattggag
      961 aacttccaaa aggtggaaaa gatcggagag ggcacgtacg gagttgtgta caaagccaga
     1021 aacaagttga cgggagaggt ggtggcgctt aagaaaatcc gcctggacac tgagactgag
     1081 ggtgtgccca gtactgccat ccgagagatc tctctgctta aggagcttaa ccatcctaat
     1141 attgtcaagc tgctggatgt cattcacaca gaaaataaac tctacctggt ttttgaattt
     1201 ctgcaccaag atctcaagaa attcatggat gcctctgctc tcactggcat tcctcttccc
     1261 ctcatcaaga gctatctgtt ccagctgctc cagggcctag ctttctgcca ttctcatcgg
     1321 gtcctccacc gagaccttaa acctcagaat ctgcttatta acacagaggg ggccatcaag
     1381 ctagcagact ttggactagc cagagctttt ggagtccctg ttcgtactta cacccatgag
     1441 gtggtgaccc tgtggtaccg agctcctgaa atcctcctgg gctgcaaata ttattccaca
     1501 gctgtggaca tctggagcct gggctgcatc tttgctgaga tggtgactcg ccgggcccta
     1561 ttccctggag attctgagat tgaccagctc ttccggatct ttcggactct ggggacccca
     1621 gatgaggtgg tgtggccagg agttacttct atgcctgatt acaagccaag tttccccaag
     1681 tgggcccggc aagattttag taaagttgta cctcccctgg atgaagatgg acggagcttg
     1741 ttatcgcaaa tgctgcacta cgaccctaac aagcggattt cggccaaggc agccctggct
     1801 caccctttct tccaggatgt gaccaagcca gtaccccatc ttcgactctg aaattctgca
     1861 gatatccagc acagtggcgg ccgctcgagc atgcatctag agggccctat tctatagtgt
     1921 cacctaaatg ctagagctcg ctgatcagcc tcgactgtgc cttctagttg ccagccatct
     1981 gttgtttgcc cctcccccgt gccttccttg accctggaag gtgccactcc cactgtcctt
     2041 tcctaataaa atgaggaaat tgcatcgcat tgtctgagta ggtgtcattc tattctgggg
     2101 ggtggggtgg ggcaggacag caagggggag gattgggaag acaatagcag gcatgctggg
     2161 gatgcggtgg gctctatggc ttctgaggcg gaaagaacca gctggggctc tagggggtat
     2221 ccccacgcgc cctgtagcgg cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg
     2281 accgctacac ttgccagcgc cctagcgccc gctcctttcg ctttcttccc ttcctttctc
     2341 gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt agggttccga
     2401 tttagtgctt tacggcacct cgaccccaaa aaacttgatt agggtgatgg ttcacgtagt
     2461 gggccatcgc cctgatagac ggtttttcgc cctttgacgt tggagtccac gttctttaat
     2521 agtggactct tgttccaaac tggaacaaca ctcaacccta tctcggtcta ttcttttgat
     2581 ttataaggga ttttgccgat ttcggcctat tggttaaaaa atgagctgat ttaacaaaaa
     2641 tttaacgcga attaattctg tggaatgtgt gtcagttagg gtgtggaaag tccccaggct
     2701 ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc aggtgtggaa
     2761 agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa
     2821 ccatagtccc gcccctaact ccgcccatcc cgcccctaac tccgcccagt tccgcccatt
     2881 ctccgcccca tggctgacta atttttttta tttatgcaga ggccgaggcc gcctctgcct
     2941 ctgagctatt ccagaagtag tgaggaggct tttttggagg cctaggcttt tgcaaaaagc
     3001 tcccgggagc ttgtatatcc attttcggat ctgatcaaga gacaggatga ggatcgtttc
     3061 gcatgattga acaagatgga ttgcacgcag gttctccggc cgcttgggtg gagaggctat
     3121 tcggctatga ctgggcacaa cagacaatcg gctgctctga tgccgccgtg ttccggctgt
     3181 cagcgcaggg gcgcccggtt ctttttgtca agaccgacct gtccggtgcc ctgaatgaac
     3241 tgcaggacga ggcagcgcgg ctatcgtggc tggccacgac gggcgttcct tgcgcagctg
     3301 tgctcgacgt tgtcactgaa gcgggaaggg actggctgct attgggcgaa gtgccggggc
     3361 aggatctcct gtcatctcac cttgctcctg ccgagaaagt atccatcatg gctgatgcaa
     3421 tgcggcggct gcatacgctt gatccggcta cctgcccatt cgaccaccaa gcgaaacatc
     3481 gcatcgagcg agcacgtact cggatggaag ccggtcttgt cgatcaggat gatctggacg
     3541 aagagcatca ggggctcgcg ccagccgaac tgttcgccag gctcaaggcg cgcatgcccg
     3601 acggcgagga tctcgtcgtg acccatggcg atgcctgctt gccgaatatc atggtggaaa
     3661 atggccgctt ttctggattc atcgactgtg gccggctggg tgtggcggac cgctatcagg
     3721 acatagcgtt ggctacccgt gatattgctg aagagcttgg cggcgaatgg gctgaccgct
     3781 tcctcgtgct ttacggtatc gccgctcccg attcgcagcg catcgccttc tatcgccttc
     3841 ttgacgagtt cttctgagcg ggactctggg gttcgaaatg accgaccaag cgacgcccaa
     3901 cctgccatca cgagatttcg attccaccgc cgccttctat gaaaggttgg gcttcggaat
     3961 cgttttccgg gacgccggct ggatgatcct ccagcgcggg gatctcatgc tggagttctt
     4021 cgcccacccc aacttgttta ttgcagctta taatggttac aaataaagca atagcatcac
     4081 aaatttcaca aataaagcat ttttttcact gcattctagt tgtggtttgt ccaaactcat
     4141 caatgtatct tatcatgtct gtataccgtc gacctctagc tagagcttgg cgtaatcatg
     4201 gtcatagctg tttcctgtgt gaaattgtta tccgctcaca attccacaca acatacgagc
     4261 cggaagcata aagtgtaaag cctggggtgc ctaatgagtg agctaactca cattaattgc
     4321 gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc attaatgaat
     4381 cggccaacgc gcggggagag gcggtttgcg tattgggcgc tcttccgctt cctcgctcac
     4441 tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta tcagctcact caaaggcggt
     4501 aatacggtta tccacagaat caggggataa cgcaggaaag aacatgtgag caaaaggcca
     4561 gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata ggctccgccc
     4621 ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc cgacaggact
     4681 ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg ttccgaccct
     4741 gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc tttctcatag
     4801 ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca
     4861 cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc ttgagtccaa
     4921 cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga ttagcagagc
     4981 gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg gctacactag
     5041 aagaacagta tttggtatct gcgctctgct gaagccagtt accttcggaa aaagagttgg
     5101 tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg tttgcaagca
     5161 gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt ctacggggtc
     5221 tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgagat tatcaaaaag
     5281 gatcttcacc tagatccttt taaattaaaa atgaagtttt aaatcaatct aaagtatata
     5341 tgagtaaact tggtctgaca gttaccaatg cttaatcagt gaggcaccta tctcagcgat
     5401 ctgtctattt cgttcatcca tagttgcctg actccccgtc gtgtagataa ctacgatacg
     5461 ggagggctta ccatctggcc ccagtgctgc aatgataccg cgagacccac gctcaccggc
     5521 tccagattta tcagcaataa accagccagc cggaagggcc gagcgcagaa gtggtcctgc
     5581 aactttatcc gcctccatcc agtctattaa ttgttgccgg gaagctagag taagtagttc
     5641 gccagttaat agtttgcgca acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc
     5701 gtcgtttggt atggcttcat tcagctccgg ttcccaacga tcaaggcgag ttacatgatc
     5761 ccccatgttg tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg tcagaagtaa
     5821 gttggccgca gtgttatcac tcatggttat ggcagcactg cataattctc ttactgtcat
     5881 gccatccgta agatgctttt ctgtgactgg tgagtactca accaagtcat tctgagaata
     5941 gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata cgggataata ccgcgccaca
     6001 tagcagaact ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa aactctcaag
     6061 gatcttaccg ctgttgagat ccagttcgat gtaacccact cgtgcaccca actgatcttc
     6121 agcatctttt actttcacca gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc
     6181 aaaaaaggga ataagggcga cacggaaatg ttgaatactc atactcttcc tttttcaata
     6241 ttattgaagc atttatcagg gttattgtct catgagcgga tacatatttg aatgtattta
     6301 gaaaaataaa caaatagggg ttccgcgcac atttccccga aaagtgccac ctgacgtc
//
