LOCUS       sequence-375410-        7110 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Overexpression of Halo-tDeg in human cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7110)
  AUTHORS   Yang LZ, Gao BQ, Huang Y, Wang Y, Yang L, Chen LL
  TITLE     Multi-color RNA imaging with CRISPR-Cas13b systems in living cells.
  JOURNAL   Cell Insight. 2022 Jun 15;1(4):100044. doi: 
            10.1016/j.cellin.2022.100044. eCollection 2022 Aug.
  PUBMED    37192858
REFERENCE   2  (bases 1 to 7110)
  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: "Cell 
            Insight."; date: "2022-06-15"; volume: "1(4):100044. doi"; pages: " 
            10.1016/j.cellin.2022.100044. eCollection 2022 Aug"
COMMENT     Sequence Label: pHAGE-Halo-tDeg
FEATURES             Location/Qualifiers
     source          1..7110
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     LTR             1..634
                     /label=3' LTR
                     /note="3' long terminal repeat (LTR) from HIV-1"
     misc_feature    681..806
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1303..1536
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             1721..1765
                     /codon_start=1
                     /product="antigenic peptide corresponding to amino acids 
                     655 to 669 of the HIV envelope protein gp41 (Lutje Hulsik 
                     et al., 2013)"
                     /label=gp41 peptide
                     /note="recognized by the 2H10 single-chain llama nanobody"
                     /translation="KNEQELLELDKWASL"
     misc_feature    2058..2175
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        2265..2476
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     CDS             2530..3414
                     /codon_start=1
                     /product="modified bacterial dehalogenase that forms 
                     covalent bonds with chloroalkane derivatives"
                     /label=HaloTag(R)
                     /translation="EIGTGFPFDPHYVEVLGERMHYVDVGPRDGTPVLFLHGNPTSSYV
                     WRNIIPHVAPTHRCIAPDLIGMGKSDKPDLGYFFDDHVRFMDAFIEALGLEEVVLVIHD
                     WGSALGFHWAKRNPERVKGIAFMEFIRPIPTWDEWPEFARETFQAFRTTDVGRKLIIDQ
                     NVFIEGTLPMGVVRPLTEVEMDHYREPFLNPVDREPLWRFPNELPIAGEPANIVALVEE
                     YMDWLHQSPVPKLLFWGTPGVLIPPAEAARLAKSLPNCKAVDIGPGLNLLQEDNPDLIG
                     SEIARWLSTLEISG"
     CDS             3421..3441
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     misc_feature    3500..4088
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3553..3573)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3971..3982)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             4163..4396
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     primer_bind     complement(4515..4533)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4601..4705
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4706..4774,4775..5566)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(4924..4943)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5737..6325
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6226..6245
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6479..6496
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    6736..6757
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(6772..6777,6778..6795,6796..6802)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6810..6826
                     /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     6815..6837
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     6834..6850
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6834..6850
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     6900..6919
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
ORIGIN
        1 tggaagggct aattcactcc caaagaagac aagatatcct tgatctgtgg atctaccaca
       61 cacaaggcta cttccctgat tagcagaact acacaccagg gccaggggtc agatatccac
      121 tgacctttgg atggtgctac aagctagtac cagttgagcc agataaggta gaagaggcca
      181 ataaaggaga gaacaccagc ttgttacacc ctgtgagcct gcatgggatg gatgacccgg
      241 agagagaagt gttagagtgg aggtttgaca gccgcctagc atttcatcac gtggcccgag
      301 agctgcatcc ggagtacttc aagaactgct gatatcgagc ttgctacaag ggactttccg
      361 ctggggactt tccagggagg cgtggcctgg gcgggactgg ggagtggcga gccctcagat
      421 cctgcatata agcagctgct ttttgcctgt actgggtctc tctggttaga ccagatctga
      481 gcctgggagc tctctggcta actagggaac ccactgctta agcctcaata aagcttgcct
      541 tgagtgcttc aagtagtgtg tgcccgtctg ttgtgtgact ctggtaacta gagatccctc
      601 agaccctttt agtcagtgtg gaaaatctct agcagtggcg cccgaacagg gacttgaaag
      661 cgaaagggaa accagaggag ctctctcgac gcaggactcg gcttgctgaa gcgcgcacgg
      721 caagaggcga ggggcggcga ctggtgagta cgccaaaaat tttgactagc ggaggctaga
      781 aggagagaga tgggtgcgag agcgtcagta ttaagcgggg gagaattaga tcgcgatggg
      841 aaaaaattcg gttaaggcca gggggaaaga aaaaatataa attaaaacat atagtatggg
      901 caagcaggga gctagaacga ttcgcagtta atcctggcct gttagaaaca tcagaaggct
      961 gtagacaaat actgggacag ctacaaccat cccttcagac aggatcagaa gaacttagat
     1021 cattatataa tacagtagca accctctatt gtgtgcatca aaggatagag ataaaagaca
     1081 ccaaggaagc tttagacaag atagaggaag agcaaaacaa aagtaagacc accgcacagc
     1141 aagcggccgg ccgctgatct tcagacctgg aggaggagat atgagggaca attggagaag
     1201 tgaattatat aaatataaag tagtaaaaat tgaaccatta ggagtagcac ccaccaaggc
     1261 aaagagaaga gtggtgcaga gagaaaaaag agcagtggga ataggagctt tgttccttgg
     1321 gttcttggga gcagcaggaa gcactatggg cgcagcgtca atgacgctga cggtacaggc
     1381 cagacaatta ttgtctggta tagtgcagca gcagaacaat ttgctgaggg ctattgaggc
     1441 gcaacagcat ctgttgcaac tcacagtctg gggcatcaag cagctccagg caagaatcct
     1501 ggctgtggaa agatacctaa aggatcaaca gctcctgggg atttggggtt gctctggaaa
     1561 actcatttgc accactgctg tgccttggaa tgctagttgg agtaataaat ctctggaaca
     1621 gatttggaat cacacgacct ggatggagtg ggacagagaa attaacaatt acacaagctt
     1681 aatacactcc ttaattgaag aatcgcaaaa ccagcaagaa aagaatgaac aagaattatt
     1741 ggaattagat aaatgggcaa gtttgtggaa ttggtttaac ataacaaatt ggctgtggta
     1801 tataaaatta ttcataatga tagtaggagg cttggtaggt ttaagaatag tttttgctgt
     1861 actttctata gtgaatagag ttaggcaggg atattcacca ttatcgtttc agacccacct
     1921 cccaaccccg aggggacccg acaggcccga aggaatagaa gaagaaggtg gagagagaga
     1981 cagagacaga tccattcgat tagtgaacgg atctcgacgg tatcgccgaa ttgacaaatg
     2041 gcagtattca tccacaattt taaaagaaaa ggggggattg gggggtacag tgcaggggaa
     2101 agaatagtag acataatagc aacagacata caaactaaag aattacaaaa acaaattaca
     2161 aaaattcaaa attttcgggt ttattacagg gacagcagag atccagtttg gactagcgat
     2221 gctagctgat gcgggccctt cgagtggctc cggtgcccgt cagtgggcag agcgcacatc
     2281 gcccacagtc cccgagaagt tggggggagg ggtcggcaat tgaaccggtg cctagagaag
     2341 gtggcgcggg gtaaactggg aaagtgatgt cgtgtactgg ctccgccttt ttcccgaggg
     2401 tgggggagaa ccgtatataa gtgcagtagt cgccgtgaac gttctttttc gcaacgggtt
     2461 tgccgccaga acacaggtgt cgtgacgcgg ttccggttaa ggtgaacgcg tgactagtgg
     2521 atgggatccg aaatcggtac tggctttcca ttcgaccccc attatgtgga agtcctgggc
     2581 gagcgcatgc actacgtcga tgttggtccg cgcgatggca cccctgtgct gttcctgcac
     2641 ggtaacccga cctcctccta cgtgtggcgc aacatcatcc cgcatgttgc accgacccat
     2701 cgctgcattg ctccagacct gatcggtatg ggcaaatccg acaaaccaga cctgggttat
     2761 ttcttcgacg accacgtccg cttcatggat gccttcatcg aagccctggg tctggaagag
     2821 gtcgtcctgg tcattcacga ctggggctcc gctctgggtt tccactgggc caagcgcaat
     2881 ccagagcgcg tcaaaggtat tgcatttatg gagttcatcc gccctatccc gacctgggac
     2941 gaatggccag aatttgcccg cgagaccttc caggccttcc gcaccaccga cgtcggccgc
     3001 aagctgatca tcgatcagaa cgtttttatc gagggtacgc tgccgatggg tgtcgtccgc
     3061 ccgctgactg aagtcgagat ggaccattac cgcgagccgt tcctgaatcc tgttgaccgc
     3121 gagccactgt ggcgcttccc aaacgagctg ccaatcgccg gtgagccagc gaacatcgtc
     3181 gcgctggtcg aagaatacat ggactggctg caccagtccc ctgtcccgaa gctgctgttc
     3241 tggggcaccc caggcgttct gatcccaccg gccgaagccg ctcgcctggc caaaagcctg
     3301 cctaactgca aggctgtgga catcggcccg ggtctgaatc tgctgcaaga agacaacccg
     3361 gacctgatcg gcagcgagat cgcgcgctgg ctgtctactc tggagatttc cggtctcgag
     3421 ccaaagaaaa agcggaaagt gagcggacct agaccaagag gaaccagagg taagggaaga
     3481 aggattcggc gcagggggta atcaacctct ggattacaaa atttgtgaaa gattgactgg
     3541 tattcttaac tatgttgctc cttttacgct atgtggatac gctgctttaa tgcctttgta
     3601 tcatgctatt gcttcccgta tggctttcat tttctcctcc ttgtataaat cctggttgct
     3661 gtctctttat gaggagttgt ggcccgttgt caggcaacgt ggcgtggtgt gcactgtgtt
     3721 tgctgacgca acccccactg gttggggcat tgccaccacc tgtcagctcc tttccgggac
     3781 tttcgctttc cccctcccta ttgccacggc ggaactcatc gccgcctgcc ttgcccgctg
     3841 ctggacaggg gctcggctgt tgggcactga caattccgtg gtgttgtcgg ggaaatcatc
     3901 gtcctttcct tggctgctcg cctgtgttgc cacctggatt ctgcgcggga cgtccttctg
     3961 ctacgtccct tcggccctca atccagcgga ccttccttcc cgcggcctgc tgccggctct
     4021 gcggcctctt ccgcgtcttc gccttcgccc tcagacgagt cggatctccc tttgggccgc
     4081 ctccccgcct gagatccttt aagaccaatg acttacaagg cagctgtaga tcttagccac
     4141 tttttaaaag aaaagggggg actggaaggg ctaattcact cccaacgaag acaagatctg
     4201 ctttttgctt gtactgggtc tctctggtta gaccagatct gagcctggga gctctctggc
     4261 taactaggga acccactgct taagcctcaa taaagcttgc cttgagtgct tcaagtagtg
     4321 tgtgcccgtc tgttgtgtga ctctggtaac tagagatccc tcagaccctt ttagtcagtg
     4381 tggaaaatct ctagcagtag tagttcatgt catcttatta ttcagtattt ataacttgca
     4441 aagaaatgaa tatcagagag tgagaggccc gggttaatta aggaaagggc tagatcattc
     4501 ttgaagacga aagggcctcg tgatacgcct atttttatag gttaatgtca tgataataat
     4561 ggtttcttag acgtcaggtg gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt
     4621 atttttctaa atacattcaa atatgtatcc gctcatgaga caataaccct gataaatgct
     4681 tcaataatat tgaaaaagga agagtatgag tattcaacat ttccgtgtcg cccttattcc
     4741 cttttttgcg gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa
     4801 agatgctgaa gatcagttgg gtgcacgagt gggttacatc gaactggatc tcaacagcgg
     4861 taagatcctt gagagttttc gccccgaaga acgttttcca atgatgagca cttttaaagt
     4921 tctgctatgt ggcgcggtat tatcccgtgt tgacgccggg caagagcaac tcggtcgccg
     4981 catacactat tctcagaatg acttggttga gtactcacca gtcacagaaa agcatcttac
     5041 ggatggcatg acagtaagag aattatgcag tgctgccata accatgagtg ataacactgc
     5101 ggccaactta cttctgacaa cgatcggagg accgaaggag ctaaccgctt ttttgcacaa
     5161 catgggggat catgtaactc gccttgatcg ttgggaaccg gagctgaatg aagccatacc
     5221 aaacgacgag cgtgacacca cgatgcctgt agcaatggca acaacgttgc gcaaactatt
     5281 aactggcgaa ctacttactc tagcttcccg gcaacaatta atagactgga tggaggcgga
     5341 taaagttgca ggaccacttc tgcgctcggc ccttccggct ggctggttta ttgctgataa
     5401 atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca gcactggggc cagatggtaa
     5461 gccctcccgt atcgtagtta tctacacgac ggggagtcag gcaactatgg atgaacgaaa
     5521 tagacagatc gctgagatag gtgcctcact gattaagcat tggtaactgt cagaccaagt
     5581 ttactcatat atactttaga ttgatttaaa acttcatttt taatttaaaa ggatctaggt
     5641 gaagatcctt tttgataatc tcatgaccaa aatcccttaa cgtgagtttt cgttccactg
     5701 agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt ttctgcgcgt
     5761 aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt tgccggatca
     5821 agagctacca actctttttc cgaaggtaac tggcttcagc agagcgcaga taccaaatac
     5881 tgttcttcta gtgtagccgt agttaggcca ccacttcaag aactctgtag caccgcctac
     5941 atacctcgct ctgctaatcc tgttaccagt ggctgctgcc agtggcgata agtcgtgtct
     6001 taccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg gctgaacggg
     6061 gggttcgtgc acacagccca gcttggagcg aacgacctac accgaactga gatacctaca
     6121 gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca ggtatccggt
     6181 aagcggcagg gtcggaacag gagagcgcac gagggagctt ccagggggaa acgcctggta
     6241 tctttatagt cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc
     6301 gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc
     6361 cttttgctgg ccttttgctc acatgttctt tcctgcgtta tcccctgatt ctgtggataa
     6421 ccgtattacc gcctttgagt gagctgatac cgctcgccgc agccgaacga ccgagcgcag
     6481 cgagtcagtg agcgaggaag cggaagagcg cccaatacgc aaaccgcctc tccccgcgcg
     6541 ttggccgatt cattaatgca gcaagctcat ggctgactaa ttttttttat ttatgcagag
     6601 gccgaggccg cctcggcctc tgagctattc cagaagtagt gaggaggctt ttttggaggc
     6661 ctaggctttt gcaaaaagct ccccgtggca cgacaggttt cccgactgga aagcgggcag
     6721 tgagcgcaac gcaattaatg tgagttagct cactcattag gcaccccagg ctttacactt
     6781 tatgcttccg gctcgtatgt tgtgtggaat tgtgagcgga taacaatttc acacaggaaa
     6841 cagctatgac atgattacga atttcacaaa taaagcattt ttttcactgc attctagttg
     6901 tggtttgtcc aaactcatca atgtatctta tcatgtctgg atcaactgga taactcaagc
     6961 taaccaaaat catcccaaac ttcccacccc ataccctatt accactgcca attacctgtg
     7021 gtttcattta ctctaaacct gtgattcctc tgaattattt tcattttaaa gaaattgtat
     7081 ttgttaaata tgtactacaa acttagtagt
//
