LOCUS       sequence-210553-        5019 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Recipient vector for expression of N-terminally Avi-tagged protein 
            of interest in chick embryos under the control of enhancer of 
            choice..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5019)
  AUTHORS   Lukoseviciute M, Ling ITC, Senanayake U, Candido-Ferreira I, Taylor 
            G, Williams RM, Sauka-Spengler T
  TITLE     Tissue-Specific In Vivo Biotin Chromatin Immunoprecipitation with 
            Sequencing in Zebrafish and Chicken.
  JOURNAL   STAR Protoc. 2020 Jul 31;1(2):100066. doi: 
            10.1016/j.xpro.2020.100066. eCollection 2020 Sep 18.
  PUBMED    33111104
REFERENCE   2  (bases 1 to 5019)
  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: "STAR 
            Protoc."; date: "2020-07-31"; volume: "1(2):100066. doi"; pages: " 
            10.1016/j.xpro.2020.100066. eCollection 2020 Sep 18"
COMMENT     Sequence Label: pTK-ENH_BsmB1-Avi-MCS-2A-Citrine
FEATURES             Location/Qualifiers
     source          1..5019
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     polyA_signal    1..49
                     /note="synthetic polyadenylation signal"
     misc_feature    63..154
                     /label=pause site
                     /note="RNA polymerase II transcriptional pause signal from 
                     the human alpha-2 globin gene"
     primer_bind     103..122
                     /label=RVprimer3
                     /note="pGL3 vector, forward primer"
     primer_bind     337..359
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     351..368
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     352..368
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     378..397
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        378..396
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(444..464)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(444..462)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(483..499)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(483..499)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(496..518)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    507..523
                     /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(531..537,538..555,556..561))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    576..597
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        765..910
                     /label=HSV TK promoter
                     /note="herpes simplex virus thymidine kinase promoter"
     intron          988..1120
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     1103..1122
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     regulatory      1174..1183
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1189..1233
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             1240..1260
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             1264..1287
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             1303..1356
                     /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             join(1372..1374,1375..1377,1378..2088)
                     /codon_start=1
                     /product="enhanced variant of YFP (Heikal et al., 2001)"
                     /label=Citrine
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLMCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1417..1438)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1678..1697)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2025..2046
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    2143..2264
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2154..2173)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2208..2227
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(2512..2529)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2683..3271)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2763..2782)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3442..4233,4234..4302))
                     /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     4065..4084
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4303..4407)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      4434..4889
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4521..4540)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4731..4752
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(join(4967..5019,1..487))
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITPSAQLTLTKGNKSWRTSRGGPVPNSPYSESYYARSLAVVL
                     QRRDWENPGVTQLNRLAAHPPFASWRNSEEARTDRPSQQLRSLNGEWDAPCSGARLPGN
                     SSSVPIDREMFWHLHLHWGQPILLVCFVSFCFDGERMLVLSIHTKNQHTDVMKIKIFYC
                     GRSKYLPYLNITYLSW"
ORIGIN
        1 aataaaatat ctttattttc attacatctg tgtgttggtt ttttgtgtga atcgatagta
       61 ctaacatacg ctctccatca aaacaaaacg aaacaaaaca aactagcaaa ataggctgtc
      121 cccagtgcaa gtgcaggtgc cagaacattt ctctatcgat aggtaccgag ctcgagttgc
      181 cagggagacg ggcgccgcta cagggcgcgt cccattcgcc attcaggctg cgcaactgtt
      241 gggaagggcg atcggtgcgg gcctcttcgc tattacgcca gctggcgaaa gggggatgtg
      301 ctgcaaggcg attaagttgg gtaacgccag ggttttccca gtcacgacgt tgtaaaacga
      361 cggccagtga gcgcgcgtaa tacgactcac tatagggcga attgggtacc gggccccccc
      421 tcgaggtcct ccagcttttg ttccctttag tgagggttaa ttgcgcgctt ggcgtaatca
      481 tggtcatagc tgtttcctgt gtgaaattgt tatccgctca caattccaca caacatacga
      541 gccggaagca taaagtgtaa agcctggggt gcctaatgag tgagctaact cacattaatt
      601 gcgttgcgct cactgcccgc tttccaccgg tcgtctcctg ttggctagcc cgggctcgag
      661 atctgcgatc taagtaagct tgctgcttca tccccgtggc ccgttgctcg cgtttgctgg
      721 cggtgtcccc ggaagaaata tatttgcatg tctttagttc tatgatgaca caaaccccgc
      781 ccagcgtctt gtcattggcg aattcgaaca cgcagatgca gtcggggcgg cgcggtccca
      841 ggtccacttc gcatattaag gtgacgcgtg tggcctcgaa caccgagcga ccctgcagcg
      901 acccgcttaa cagcgtcaac agcgtgccgc agatcccaag cttgcggccg cttaagctgc
      961 agaagttggt cgtgaggcac tgggcaggta agtatcaagg ttacaagaca ggtttaagga
     1021 gaccaataga aactgggctt gtcgagacag agaagactct tgcgtttctg ataggcacct
     1081 attggtctta ctgacatcca ctttgccttt ctctccacag gtgtccactc ccaggttcaa
     1141 tacagctctt aagcggccgc aagctagctt gccgccacca tggctggtgg cctgaatgac
     1201 atctttgagg cccagaagat cgagtggcat gagggaggag agaacctgta cttccaggga
     1261 ggagactaca aggacgacga cgacaaggat atcggatcag gagagggcag gggaagtctt
     1321 ctaacatgcg gggacgtgga ggaaaatccc gggcccggcg gatccggagg catggtgagc
     1381 aagggcgagg agctgttcac cggggtggtg cccatcctgg tcgagctgga cggcgacgta
     1441 aacggccaca agttcagcgt gtccggcgag ggcgagggcg atgccaccta cggcaagctg
     1501 accctgaagt tcatctgcac caccggcaag ctgcccgtgc cctggcccac cctcgtgacc
     1561 accttcggct acggcctgat gtgcttcgcc cgctaccccg accacatgaa gcagcacgac
     1621 ttcttcaagt ccgccatgcc cgaaggctac gtccaggagc gcaccatctt cttcaaggac
     1681 gacggcaact acaagacccg cgccgaggtg aagttcgagg gcgacaccct ggtgaaccgc
     1741 atcgagctga agggcatcga cttcaaggag gacggcaaca tcctggggca caagctggag
     1801 tacaactaca acagccacaa cgtctatatc atggccgaca agcagaagaa cggcatcaag
     1861 gtgaacttca agatccgcca caacatcgag gacggcagcg tgcagctcgc cgaccactac
     1921 cagcagaaca cccccatcgg cgacggcccc gtgctgctgc ccgacaacca ctacctgagc
     1981 taccagtccg ccctgagcaa agaccccaac gagaagcgcg atcacatggt cctgctggag
     2041 ttcgtgaccg ccgccgggat cactctcggc atggacgagc tgtacaagta aagcggccgc
     2101 gactctagag tcggggcggc cggccgcttc gagcagacat gataagatac attgatgagt
     2161 ttggacaaac cacaactaga atgcagtgaa aaaaatgctt tatttgtgaa atttgtgatg
     2221 ctattgcttt atttgtaacc attataagct gcaataaaca agttaacaac aacaattgca
     2281 ttcattttat gtttcaggtt cagggggagg tgtgggaggt tttttaaagc aagtaaaacc
     2341 tctacaaatg tggtaaaatc gataaggatc cgtcgaccga tgcccttgag agccttcaac
     2401 ccagtcagct ccttccggtg ggcgcggggc atgactatcg tcgccgcact tatgactgtc
     2461 ttctttatca tgcaactcgt aggacaggtg ccggcagcgc tcttccgctt cctcgctcac
     2521 tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta tcagctcact caaaggcggt
     2581 aatacggtta tccacagaat caggggataa cgcaggaaag aacatgtgag caaaaggcca
     2641 gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata ggctccgccc
     2701 ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc cgacaggact
     2761 ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg ttccgaccct
     2821 gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc tttctcatag
     2881 ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca
     2941 cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc ttgagtccaa
     3001 cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga ttagcagagc
     3061 gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg gctacactag
     3121 aagaacagta tttggtatct gcgctctgct gaagccagtt accttcggaa aaagagttgg
     3181 tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg tttgcaagca
     3241 gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt ctacggggtc
     3301 tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgagat tatcaaaaag
     3361 gatcttcacc tagatccttt taaattaaaa atgaagtttt aaatcaatct aaagtatata
     3421 tgagtaaact tggtctgaca gttaccaatg cttaatcagt gaggcaccta tctcagcgat
     3481 ctgtctattt cgttcatcca tagttgcctg actccccgtc gtgtagataa ctacgatacg
     3541 ggagggctta ccatctggcc ccagtgctgc aatgataccg cgagacccac gctcaccggc
     3601 tccagattta tcagcaataa accagccagc cggaagggcc gagcgcagaa gtggtcctgc
     3661 aactttatcc gcctccatcc agtctattaa ttgttgccgg gaagctagag taagtagttc
     3721 gccagttaat agtttgcgca acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc
     3781 gtcgtttggt atggcttcat tcagctccgg ttcccaacga tcaaggcgag ttacatgatc
     3841 ccccatgttg tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg tcagaagtaa
     3901 gttggccgca gtgttatcac tcatggttat ggcagcactg cataattctc ttactgtcat
     3961 gccatccgta agatgctttt ctgtgactgg tgagtactca accaagtcat tctgagaata
     4021 gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata cgggataata ccgcgccaca
     4081 tagcagaact ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa aactctcaag
     4141 gatcttaccg ctgttgagat ccagttcgat gtaacccact cgtgcaccca actgatcttc
     4201 agcatctttt actttcacca gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc
     4261 aaaaaaggga ataagggcga cacggaaatg ttgaatactc atactcttcc tttttcaata
     4321 ttattgaagc atttatcagg gttattgtct catgagcgga tacatatttg aatgtattta
     4381 gaaaaataaa caaatagggg ttccgcgcac atttccccga aaagtgccac ctgacgcgcc
     4441 ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga ccgctacact
     4501 tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct tcctttctcg ccacgttcgc
     4561 cggctttccc cgtcaagctc taaatcgggg gctcccttta gggttccgat ttagtgcttt
     4621 acggcacctc gaccccaaaa aacttgatta gggtgatggt tcacgtagtg ggccatcgcc
     4681 ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg ttctttaata gtggactctt
     4741 gttccaaact ggaacaacac tcaaccctat ctcggtctat tcttttgatt tataagggat
     4801 tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa
     4861 ttttaacaaa atattaacgc ttacaatttg ccattcgcca ttcaggctgc gcaactgttg
     4921 ggaagggcga tcggtgcggg cctcttcgct attacgccag cccaagctac catgataagt
     4981 aagtaatatt aaggtacggg aggtacttgg agcggccgc
//
