LOCUS       sequence-368304-        6233 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  CAG promoter-driven expression of BioID2-fused protein (C terminus).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6233)
  AUTHORS   Oura S, Ninomiya A, Sugihara F, Matzuk MM, Ikawa M
  TITLE     Proximity-dependent biotin labeling in testicular germ cells 
            identified TESMIN-associated proteins.
  JOURNAL   Sci Rep. 2022 Dec 23;12(1):22198. doi: 10.1038/s41598-022-26501-7.
  PUBMED    36564444
REFERENCE   2  (bases 1 to 6233)
  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: "Sci Rep."; 
            date: "2022-12-23"; volume: "12(1):22198. doi"; pages: " 
            10.1038/s41598-022-26501-7"
COMMENT     Sequence Label: pCAG-MCS-TurboID-3xFLAG
FEATURES             Location/Qualifiers
     source          1..6233
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        17..396
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     intron          675..1689
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1615..1637
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1697..1716
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     CDS             1789..2745
                     /codon_start=1
                     /product="mutant of the E. coli biotin ligase BioID 
                     engineered for improved catalytic efficiency (Branon et 
                     al., 2018)"
                     /label=TurboID
                     /translation="KDNTVPLKLIALLANGEFHSGEQLGETLGMSRAAINKHIQTLRDW
                     GVDVFTVPGKGYSLPEPIPLLNAKQILGQLDGGSVAVLPVVDSTNQYLLDRIGELKSGD
                     ACIAEYQQAGRGSRGRKWFSPFGANLYLSMFWRLKRGPAAIGLGPVIGIVMAEALRKLG
                     ADKVRVKWPNDLYLQDRKLAGILVELAGITGDAAQIVIGAGINVAMRRVEESVVNQGWI
                     TLQEAGINLDRNTLAATLIRELRAALELFEQEGLAPYLPRWEKLDNFINRPVKLIIGDK
                     EIFGISRGIDKQGALLLEQDGVIKPWMGGEISLRSAEK"
     CDS             2761..2826
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     primer_bind     complement(2913..2932)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    2978..3033
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(3032..3051)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(3388..3407)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(3389..3407)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3417..3434)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3417..3433)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3426..3448)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      3575..4030
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3662..3681)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3872..3893
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        4056..4160
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4161..4229,4230..5021)
                     /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"
     rep_origin      5192..5780
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5681..5700
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5934..5951
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    6068..6089
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(6104..6109,6110..6127,6128..6134)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6142..6158
                     /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     6147..6169
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     6166..6182
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6166..6182
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     6201..6221
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        6203..6221
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
ORIGIN
        1 gagctcatcg atactcgaca ttgattattg actagttatt aatagtaatc aattacgggg
       61 tcattagttc atagcccata tatggagttc cgcgttacat aacttacggt aaatggcccg
      121 cctggctgac cgcccaacga cccccgccca ttgacgtcaa taatgacgta tgttcccata
      181 gtaacgccaa tagggacttt ccattgacgt caatgggtgg agtatttacg gtaaactgcc
      241 cacttggcag tacatcaagt gtatcatatg ccaagtacgc cccctattga cgtcaatgac
      301 ggtaaatggc ccgcctggca ttatgcccag tacatgacct tatgggactt tcctacttgg
      361 cagtacatct acgtattagt catcgctatt accatggtcg aggtgagccc cacgttctgc
      421 ttcactctcc ccatctcccc cccctcccca cccccaattt tgtatttatt tattttttaa
      481 ttattttgtg cagcgatggg ggcggggggg gggggggnsg cgcgccrggs ggggsggggs
      541 ggggsgrggg ggggggsggg gsgrggcggr gaggkgcggc ggcagccaat cagagcggcg
      601 cgctccgaaa gtttcctttt atggcgaggc ggcggcggcg gcggccctat aaaaagcgaa
      661 gcgcgcggcg ggcgggagtc gctgcgcgct gccttcgccc cgtgccccgc tccgccgccg
      721 cctcgcgccg cccgccccgg ctctgactga ccgcgttact cccacaggtg agcgggcggg
      781 acggcccttc tcctccgggc tgtaattagc gcttggttta atgacggctt gtttcttttc
      841 tgtggctgcg tgaaagcctt gaggggctcc gggagggccc tttgtgcggg gggagcggct
      901 cggggggtgc gtgcgtgtgt gtgtgcgtgg ggagcgccgc gtgcggctcc gcgctgcccg
      961 gcggctgtga gcgctgcggg cgcggcgcgg ggctttgtgc gctccgcagt gtgcgcgagg
     1021 ggagcgcggc cgggggcggt gccccgcggt gcgggggggg ctgcgagggg aacaaaggct
     1081 gcgtgcgggg tgtgtgcgtg ggggggtgag cagggggtgt gggcgcgtcg gtcgggctgc
     1141 aaccccccct gcacccccct ccccgagttg ctgagcacgg cccggcttcg ggtgcggggc
     1201 tccgtacggg gcgtggcgcg gggctcgccg tgccgggcgg ggggtggcgg caggtggggg
     1261 tgccgggcgg ggcggggccg cctcgggccg gggrgggctc gggggagggg cgcggcggcc
     1321 cccggagcgc cggcggctgt cgaggcgcgg cgagccgcag ccattgcctt ttatggtaat
     1381 cgtgcgagag ggcgcaggga cttcctttgt cccaaatctg tgcggagccg aaatctggga
     1441 ggcgccgccg caccccctct agcgggcgcg gggcgaagcg gtgcggcgcc ggcaggaagg
     1501 aaatgggcgg ggagggcctt cgtgcgtcgc cgcgccgccg tccccttctc cctctccagc
     1561 ctcggggctg tccgcggggg gacggctgcc ttcggggggg acggggcagg gcggggttcg
     1621 gcttctggcg tgtgaccggc ggctctagcc tctgctaacc atgttcatgc cttcttcttt
     1681 ttcctacagc tcctgggcaa cgtgctggtt attgtgctgt ctcatcattt tggcaaatct
     1741 agaggatccc tgcaggaatt cgatatcaag cttgtcgacg cggccgcgaa agacaatact
     1801 gtgcctctga agctgatcgc tctcctggct aatggcgagt tccatagtgg cgaacagctg
     1861 ggagaaaccc tgggcatgtc cagggccgct atcaacaagc acattcagac tctgcgcgac
     1921 tggggcgtgg acgtgttcac cgtgcccgga aagggctact ctctgcccga gcctatcccg
     1981 ctgctgaacg ctaaacagat tctgggacag ctggacggcg ggagcgtggc agtcctgcct
     2041 gtggtcgact ccaccaatca gtacctgctg gatcgaatcg gcgagctgaa gagtggggat
     2101 gcttgcattg cagaatatca gcaggcaggg agaggaagca gagggaggaa atggttctct
     2161 ccttttggag ctaacctgta cctgagtatg ttttggcgcc tgaagcgggg accagcagca
     2221 atcggcctgg gcccggtcat cggaattgtc atggcagaag cgctgcgaaa gctgggagca
     2281 gacaaggtgc gagtcaaatg gcccaatgac ctgtatctgc aggatagaaa gctggcaggc
     2341 atcctggtgg agctggccgg aataacaggc gatgctgcac agatcgtcat tggcgccggg
     2401 attaacgtgg ctatgaggcg cgtggaggaa agcgtggtca atcagggctg gatcacactg
     2461 caggaagcag ggattaacct ggacaggaat actctggccg ctacgctgat ccgagagctg
     2521 cgggcagccc tggaactgtt cgagcaggaa ggcctggctc catatctgcc acggtgggag
     2581 aagctggata acttcatcaa tagacccgtg aagctgatca ttggggacaa agagattttc
     2641 gggattagcc gggggattga taaacaggga gccctgctgc tggaacagga cggagttatc
     2701 aaaccctgga tgggcggaga aatcagtctg cggtctgccg aaaaggctag cgggggcggg
     2761 gactacaaag accatgacgg tgattataaa gatcatgaca ttgattacaa ggatgacgat
     2821 gacaagtagc tcgagactcc tcaggtgcag gctgcctatc agaaggtggt ggctggtgtg
     2881 gccaatgccc tggctcacaa ataccactga gatctttttc cctctgccaa aaattatggg
     2941 gacatcatga agccccttga gcatctgact tctggctaat aaaggaaatt tattttcatt
     3001 gcaatagtgt gttggaattt tttgtgtctc tcactcggaa ggacatatgg gagggcaaat
     3061 catttaaaac atcagaatga gtatttggtt tagagtttgg caacatatgc ccatatgctg
     3121 gctgccatga acaaaggttg gctataaaga ggtcatcagt atatgaaaca gccccctgct
     3181 gtccattcct tattccatag aaaagccttg acttgaggtt agattttttt tatattttgt
     3241 tttgtgttat ttttttcttt aacatcccta aaattttcct tacatgtttt actagccaga
     3301 tttttcctcc tctcctgact actcccagtc atagctgtcc ctcttctctt atgaagatcc
     3361 ctcgacttaa ttaaggtacc caattcgccc tatagtgagt cgtattacgc gcgctcactg
     3421 gccgtcgttt tacaacgtcg tgactgggaa aaccctggcg ttacccaact taatcgcctt
     3481 gcagcacatc cccctttcgc cagctggcgt aatagcgaag aggcccgcac cgatcgccct
     3541 tcccaacagt tgcgcagcct gaatggcgaa tgggacgcgc cctgtagcgg cgcattaagc
     3601 gcggcgggtg tggtggttac gcgcagcgtg accgctacac ttgccagcgc cctagcgccc
     3661 gctcctttcg ctttcttccc ttcctttctc gccacgttcg ccggctttcc ccgtcaagct
     3721 ctaaatcggg ggctcccttt agggttccga tttagtgctt tacggcacct cgaccccaaa
     3781 aaacttgatt agggtgatgg ttcacgtagt gggccatcgc cctgatagac ggtttttcgc
     3841 cctttgacgt tggagtccac gttctttaat agtggactct tgttccaaac tggaacaaca
     3901 ctcaacccta tctcggtcta ttcttttgat ttataaggga ttttgccgat ttcggcctat
     3961 tggttaaaaa atgagctgat ttaacaaaaa tttaacgcga attttaacaa aatattaacg
     4021 cttacaattt aggtggcact tttcggggaa atgtgcgcgg aacccctatt tgtttatttt
     4081 tctaaataca ttcaaatatg tatccgctca tgagacaata accctgataa atgcttcaat
     4141 aatattgaaa aaggaagagt atgagtattc aacatttccg tgtcgccctt attccctttt
     4201 ttgcggcatt ttgccttcct gtttttgctc acccagaaac gctggtgaaa gtaaaagatg
     4261 ctgaagatca gttgggtgca cgagtgggtt acatcgaact ggatctcaac agcggtaaga
     4321 tccttgagag ttttcgcccc gaagaacgtt ttccaatgat gagcactttt aaagttctgc
     4381 tatgtggcgc ggtattatcc cgtattgacg ccgggcaaga gcaactcggt cgccgcatac
     4441 actattctca gaatgacttg gttgagtact caccagtcac agaaaagcat cttacggatg
     4501 gcatgacagt aagagaatta tgcagtgctg ccataaccat gagtgataac actgcggcca
     4561 acttacttct gacaacgatc ggaggaccga aggagctaac cgcttttttg cacaacatgg
     4621 gggatcatgt aactcgcctt gatcgttggg aaccggagct gaatgaagcc ataccaaacg
     4681 acgagcgtga caccacgatg cctgtagcaa tggcaacaac gttgcgcaaa ctattaactg
     4741 gcgaactact tactctagct tcccggcaac aattaataga ctggatggag gcggataaag
     4801 ttgcaggacc acttctgcgc tcggcccttc cggctggctg gtttattgct gataaatctg
     4861 gagccggtga gcgtgggtct cgcggtatca ttgcagcact ggggccagat ggtaagccct
     4921 cccgtatcgt agttatctac acgacgggga gtcaggcaac tatggatgaa cgaaatagac
     4981 agatcgctga gataggtgcc tcactgatta agcattggta actgtcagac caagtttact
     5041 catatatact ttagattgat ttaaaacttc atttttaatt taaaaggatc taggtgaaga
     5101 tcctttttga taatctcatg accaaaatcc cttaacgtga gttttcgttc cactgagcgt
     5161 cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct
     5221 gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc
     5281 taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgttc
     5341 ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc
     5401 tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg
     5461 ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt
     5521 cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg
     5581 agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg
     5641 gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc tggtatcttt
     5701 atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag
     5761 gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc ctggcctttt
     5821 gctggccttt tgctcacatg ttctttcctg cgttatcccc tgattctgtg gataaccgta
     5881 ttaccgcctt tgagtgagct gataccgctc gccgcagccg aacgaccgag cgcagcgagt
     5941 cagtgagcga ggaagcggaa gagcgcccaa tacgcaaacc gcctctcccc gcgcgttggc
     6001 cgattcatta atgcagctgg cacgacaggt ttcccgactg gaaagcgggc agtgagcgca
     6061 acgcaattaa tgtgagttag ctcactcatt aggcacccca ggctttacac tttatgcttc
     6121 cggctcgtat gttgtgtgga attgtgagcg gataacaatt tcacacagga aacagctatg
     6181 accatgatta cgccaagcgc gcaattaacc ctcactaaag ggaacaaaag ctg
//
