LOCUS       sequence-250990-        7052 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  HDR (donor) Plasmid for Inserting AID-GFP-2A-Puro into the 3' end of
            the Mouse Hp1a Gene.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7052)
  TITLE     Plasmids for generating AID/Degron Cell Lines in Human and Mouse
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 7052)
  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            "Unpublished"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: Mouse 3' Hp1a AID GFP PuroR
FEATURES             Location/Qualifiers
     source          1..7052
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     1..17
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     27..46
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        27..45
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     70..86
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     71..87
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             1144..1824
                     /codon_start=1
                     /gene="Arabidopsis thaliana AXR3"
                     /product="auxin-responsive IAA17 protein"
                     /label=auxin-inducible degron
                     /note="acts as an auxin-inducible degron (AID) in the 
                     presence of TIR1"
                     /translation="GSVELNLRETELCLGLPGGDTVAPVTGNKRGFSETVDLKLNLNNE
                     PANKEGSTTHDVVTFDSKEKSACPKDPAKPPAKAQVVGWPPVRSYRKNVMVSCQKSSGG
                     PEAAAFVKVSMDGAPYLRKIDLRMYKSYDELSNALSNMFSSFTMGKHGGEEGMIDFMNE
                     RKLMDLVNSWDYVPSYEDKDGDWMLVGDVPWPMFVDTCKRLRLMKGSDAIGLAPRAMEK
                     CKSRA"
     CDS             join(1831..1833,1834..2544)
                     /codon_start=1
                     /product="GFP variant that folds robustly even when fused 
                     to poorly folded proteins (Pedelacq et al., 2006)"
                     /label=superfolder GFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTITFKDDG
                     TYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIKA
                     NFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     primer_bind     complement(1873..1894)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     2481..2502
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             2554..2607
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             2614..3216
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MATEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIE
                     RVTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGSRMAELSGSRL
                     AAQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLET
                     SAPRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     3113..3133
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     primer_bind     complement(4265..4285)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(4265..4283)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(4304..4320)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4304..4320)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4317..4339)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4328..4344
                     /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(4352..4358,4359..4376,4377..4382))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4397..4418
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4535..4552)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4706..5294)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4786..4805)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5465..6256,6257..6325))
                     /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     6088..6107
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6326..6430)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(6456..6911)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6593..6614)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6805..6824
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     join(7038..7052,1..8)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     join(7052,1..17)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
ORIGIN
        1 gtaaaacgac ggccagtgag cgcgcgtaat acgactcact atagggcgaa ttgggtaccg
       61 ggccccccct cgaggtcgac ggtatcgata agcttgatat cgaattcctg cagcccgggt
      121 attgtgttat aataaccata tttgggtaaa aaaatttggg gtagaaaaat tgtattacta
      181 ctacctaaat atctatttgt atagaactaa atcatttatg taagatcatg agtctatgac
      241 tacgggataa gagctaagta aagcttccat cactgagggt cagggtcacc gaatgataga
      301 gaaattaatt tccacaaaag gcatcaaact taatgctaaa taaagcataa attactttat
      361 cagagatagt gacttataga agaaagtgga atacgctcat ccttctgctg tcctttcctg
      421 acagggtctc atgtagctca ggctgtcctt gaactcaata aggagtccag aatattaaat
      481 tcctatcccc atctccctag gactggcttt ttcctccccc tatttccttt tcttttctcc
      541 cttcccattt tcttttaaga tttatttctt atgtcattaa atgctgcata tatgtgttct
      601 gcctgaatat ataaatgtgc accatgtgta tgcagtgctt gcagaggcat tggatcccct
      661 ggaactaaca ttacaggcag tttgtgtggt accatgtagg tgctggaaac tgagcccggg
      721 tcctctgtaa gaacagccat ctcttcagct ctttctttct tcctctgtaa gaacagccat
      781 ctcttcagct ctttctttct tgtttatttt ttgatactga tgatcaaata gagaagatgt
      841 accactgagt tacacctcca gccctgtaaa gaccctttct actttgagat cgaaccagct
      901 aacctcagac agtgacactg acagattgcc tcagtttcat gtctcctaag tcttacaccc
      961 tttctttcca ggaaagacac agatgaagct gacctggttc ttgcaaaaga agctaacgtg
     1021 aagtgtccac agattgtgat agcattttat gaagagagac tgacgtggca cgcatatcca
     1081 gaggatgcgg aaaacaaaga aaaagaaagc gcgaagagcg gatccggagg gtctggaggc
     1141 tctggcagcg tggaactgaa cctgcgcgag acagagctgt gcctgggact gcctggcggc
     1201 gatacagtgg ctcctgtgac cggcaacaag agaggcttca gcgagacagt ggacctgaag
     1261 ctgaacctga acaacgagcc cgccaacaaa gagggcagca ccacccacga cgtcgtgacc
     1321 ttcgacagca aagagaagtc cgcctgcccc aaggaccccg ccaagcctcc tgctaaagct
     1381 caggtcgtgg gatggcctcc cgtgcggagc tacagaaaga acgtgatggt gtcctgccag
     1441 aagtcctctg gcggccctga ggctgccgcc ttcgtgaaag tgtctatgga tggcgcccct
     1501 tacctgagaa agatcgacct gagaatgtac aagagctacg acgagctgag caacgccctg
     1561 agcaacatgt tcagcagctt cacaatgggc aagcacggcg gcgaggaagg catgatcgac
     1621 ttcatgaacg agcggaagct gatggacctc gtgaacagct gggactacgt gcccagctac
     1681 gaggacaagg acggcgactg gatgctcgtg ggagatgtgc cctggcccat gttcgtggac
     1741 acatgcaaga gactgagact gatgaagggc agcgacgcca tcggcctggc ccccagagct
     1801 atggaaaagt gcaagagcag agccgctagc gtgagcaagg gcgaggagct gttcaccggg
     1861 gtggtgccca tcctggtcga gctggacggc gacgtaaacg gccacaagtt cagcgtgcgc
     1921 ggcgagggcg agggcgatgc caccaacggc aagctgaccc tgaagttcat ctgcaccacc
     1981 ggcaagctgc ccgtgccctg gcccaccctc gtgaccaccc tgacctacgg cgtgcagtgc
     2041 ttcagccgct accccgacca catgaagcag cacgacttct tcaagtccgc catgcccgaa
     2101 ggctacgtcc aggagcgcac catcaccttc aaggacgacg gcacctacaa gacccgcgcc
     2161 gaggtgaagt tcgagggcga caccctggtg aaccgcatcg agctgaaggg catcgacttc
     2221 aaggaggacg gcaacatcct ggggcacaag ctggagtaca acttcaacag ccacaacgtc
     2281 tatatcaccg ccgacaagca gaagaacggc atcaaggcca acttcaagat ccgccacaac
     2341 gtggaggacg gcagcgtgca gctcgccgac cactaccagc agaacacccc catcggcgac
     2401 ggccccgtgc tgctgcccga caaccactac ctgagcaccc agtccaagct gagcaaagac
     2461 cccaacgaga agcgcgatca catggtcctg ctggagttcg tgaccgccgc cgggatcact
     2521 cacggcatgg acgagctgta caagggatct ggagagggaa gaggaagcct gctgacctgt
     2581 ggagatgtgg aagagaatcc tggacctggt accatggcca ccgagtacaa gcccacggtg
     2641 cgcctcgcca cccgcgacga cgtcccccgg gccgtacgca ccctcgccgc cgcgttcgcc
     2701 gactaccccg ccacgcgcca caccgtggac ccggaccgcc acatcgagcg ggtcaccgag
     2761 ctgcaagaac tcttcctcac gcgcgtcggg ctcgacatcg gcaaggtgtg ggtcgcggac
     2821 gacggcgccg cggtggcggt ctggaccacg ccggagagcg tcgaagcggg ggcggtgttc
     2881 gccgagatcg gctcgcgcat ggccgagttg agcggttccc ggctggccgc gcagcaacag
     2941 atggaaggcc tcctggcgcc gcaccggccc aaggagcccg cgtggttcct ggccaccgtc
     3001 ggcgtctcgc ccgaccacca gggcaagggt ctgggcagcg ccgtcgtgct ccccggagtg
     3061 gaggcggccg agcgcgctgg ggtgcccgcc ttcctggaga cctccgcgcc ccgcaacctc
     3121 cccttctacg agcggctcgg cttcaccgtc accgccgacg tcgaggtgcc cgaaggaccg
     3181 cgcacctggt gcatgacccg caagcccggt gcctgataaa ggaggaggca gtctctgtcg
     3241 tttctctttg tatataatac ctttactttc ctgcctcctc tctctcctcc cacccctttc
     3301 catcctaaac gcatccataa accactgcgc tcatcactgt gctccacggg ggagatgctg
     3361 gtattggttc tcctgtaata taactgatga tctgattcct aagtgtagct ctgtgaacgt
     3421 caggcattac gtcattcaca tgctgtgtgg ttcccatagt gttctctttc accctgatct
     3481 tttccttctg ttcccctgta actttttaat caccttagag tcatgatctt tggggttggt
     3541 tgctttggga gattttatta tgacgatgac ttcagtttct tgattgttat tagattatat
     3601 tgatgaccag ctggcctttg ttttgacata ctggcataga aaggatattg tccatctatt
     3661 tagaaaacta ttaagggcaa cctacctgtt gtgtcttttc cgatctcatt tagctctgcc
     3721 aggagaatac agggtccctg gtgtggtctt gggccgctcc tgactatcct cacataccct
     3781 cccataacaa ttcagttctc ttagggcttg aaacttcaca tttaaatctg tcaaggcgct
     3841 ctgtttagcc ctcacacttt cttggccttg ttactttcaa tccctgattc tgcttctgta
     3901 aagtggtatc aatttgttat taaatgacaa gattaactgt ggagtctgta agctgtagtg
     3961 caggaagata ataggatggg gctggataca gttatcacgt ggtgtgtgtg tattctttcc
     4021 tttatccctc atatttagac tgtatttatg taggtgtgag tgattgcctg gtgcttgctt
     4081 gtgccaaggt gctgggcacc ctctaaccct gccaattttt atggcctccc aaagcattcc
     4141 tgtaccaaag aggcatcagc agatccatgc cattcccttc cgtgccatta ttttaagttg
     4201 ggaatttttg gagaggacta gttctagagc ggccgccacc gcggtggagc tccagctttt
     4261 gttcccttta gtgagggtta attgcgcgct tggcgtaatc atggtcatag ctgtttcctg
     4321 tgtgaaattg ttatccgctc acaattccac acaacatacg agccggaagc ataaagtgta
     4381 aagcctgggg tgcctaatga gtgagctaac tcacattaat tgcgttgcgc tcactgcccg
     4441 ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga
     4501 gaggcggttt gcgtattggg cgctcttccg cttcctcgct cactgactcg ctgcgctcgg
     4561 tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg ttatccacag
     4621 aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc
     4681 gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca
     4741 aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt
     4801 ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc
     4861 tgtccgcctt tctccattcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc
     4921 tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc
     4981 ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact
     5041 tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg
     5101 ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca gtatttggta
     5161 tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca
     5221 aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa
     5281 aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg
     5341 aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc
     5401 ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa acttggtctg
     5461 acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat
     5521 ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc ttaccatctg
     5581 gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat ttatcagcaa
     5641 taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta tccgcctcca
     5701 tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt aatagtttgc
     5761 gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt
     5821 cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa
     5881 aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc gcagtgttat
     5941 cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc gtaagatgct
     6001 tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg cggcgaccga
     6061 gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga actttaaaag
     6121 tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta ccgctgttga
     6181 gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct tttactttca
     6241 ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg
     6301 cgacacggaa atgttgaata ctcatactct tcctttttca atattattga agcatttatc
     6361 agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag
     6421 gggttccgcg cacatttccc cgaaaagtgc cacctaaatt gtaagcgtta atattttgtt
     6481 aaaattcgcg ttaaattttt gttaaatcag ctcatttttt aaccaatagg ccgaaatcgg
     6541 caaaatccct tataaatcaa aagaatagac cgagataggg ttgagtgttg ttccagtttg
     6601 gaacaagagt ccactattaa agaacgtgga ctccaacgtc aaagggcgaa aaaccgtcta
     6661 tcagggcgat ggcccactac gtgaaccatc accctaatca agttttttgg ggtcgaggtg
     6721 ccgtaaagca ctaaatcgga accctaaagg gagcccccga tttagagctt gacggggaaa
     6781 gccggcgaac gtggcgagaa aggaagggaa gaaagcgaaa ggagcgggcg ctagggcgct
     6841 ggcaagtgta gcggtcacgc tgcgcgtaac caccacaccc gccgcgctta atgcgccgct
     6901 acagggcgcg tcccattcgc cattcaggct gcgcaactgt tgggaagggc gatcggtgcg
     6961 ggcctcttcg ctattacgcc agctggcgaa agggggatgt gctgcaaggc gattaagttg
     7021 ggtaacgcca gggttttccc agtcacgacg tt
//
