LOCUS       Exported                7739 bp ds-DNA     circular SYN 03-SEP-2022
DEFINITION  .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  recombinant plasmid
REFERENCE   1  (bases 1 to 7739)
  AUTHORS   Triple Threat
  TITLE     Direct Submission
  JOURNAL   Exported Monday, Sep 12, 2022 from SnapGene 5.0.8
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..7739
                     /organism="recombinant plasmid"
                     /mol_type="other DNA"
     enhancer        4..383
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
                     /note="color: #ffffff"
     promoter        386..661
                     /label=chicken beta-actin promoter
                     /note="color: #ffffff; direction: RIGHT"
     intron          662..1670
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
                     /note="color: #------"
     primer_bind     1599..1619
                     /label=CAG-F
                     /note="CAG-F"
                     /note="color: black; sequence: GGCTTCTGGCGTGTGACCGGC"
     primer_bind     1705..1764
                     /label=Hg-011-CSTN2-F1L
                     /note="color: black; sequence: 
                     ATCATTTTGGCAAAGGCCACCATGTACCCATACGATGTTCCAGATTACGCTGGCTATCC
                     C"
     misc_feature    1705..1719
                     /label=recombination fragment front
                     /note="color: #00ff00; direction: RIGHT"
     misc_feature    1720..1728
                     /label=KOZAK fragment
                     /note="color: #ff9900; direction: RIGHT"
     CDS             1726..4683
                     /codon_start=1
                     /label=CSTN2
                     /note="color: #993366"
                     /translation="MYPYDVPDYAGYPYDVPDYAGSYPYDVPDYALPGRLCWVPLLLAL
                     GVGSGSGGGGDSRQRRLLAAKVNKHKPWIETSYHGVITENNDTVILDPPLVALDKDAPV
                     PFAGEICAFKIHGQELPFEAVVLNKTSGEGRLRAKSPIDCELQKEYTFIIQAYDCGAGP
                     HETAWKKSHKAVVHIQVKDVNEFAPTFKEPAYKAVVTEGKIYDSILQVEAIDEDCSPQY
                     SQICNYEIVTTDVPFAIDRNGNIRNTEKLSYDKQHQYEILVTAYDCGQKPAAQDTLVQV
                     DVKPVCKPGWQDWTKRIEYQPGSGSMPLFPSIHLETCDGAVSSLQIVTELQTNYIGKGC
                     DRETYSEKSLQKLCGASSGIIDLLPSPSAATNWTAGLLVDSSEMIFKFDGRQGAKVPDG
                     IVPKNLTDQFTITMWMKHGPSPGVRAEKETILCNSDKTEMNRHHYALYVHNCRLVFLLR
                     KDFDQADTFRPAEFHWKLDQICDKEWHYYVINVEFPVVTLYMDGATYEPYLVTNDWPIH
                     PSHIAMQLTVGACWQGGEVTKPQFAQFFHGSLASLTIRPGKMESQKVISCLQACKEGLD
                     INSLESLGQGIKYHFNPSQSILVMEGDDIGNINRALQKVSYINSRQFPTAGVRRLKVSS
                     KVQCFGEDVCISIPEVDAYVMVLQAIEPRITLRGTDHFWRPAAQFESARGVTLFPDIKI
                     VSTFAKTEAPGDVKTTDPKSEVLEEMLHNLDFCDILVIGGDLDPRQECLELNHSELHQR
                     HLDATNSTAGYSIYGVGSMSRYEQVLHHIRYRNWRPASLEARRFRIKCSELNGRYTSNE
                     FNLEVSILHEDQVSDKEHVNHLIVQPPFLQSVHHPESRSSIQHSSVVPSIATVVIIISV
                     CMLVFVVAMGVYRVRIAHQHFIQETEAAKESEMDWDDSALTITVNPMEKHEGPGHGEDE
                     TEGEEEEEAEEEMSSSSGSDDSEEEEEEEGMGRGRHGQNGARQAQLEWDDSTLPY"
     misc_feature    1726..1728
                     /label=ATG
                     /note="color: #ff0000; direction: RIGHT"
     CDS             1729..1818
                     /codon_start=1
                     /product="three tandem HA epitope tags"
                     /label=3xHA
                     /note="color: #cc99b2"
                     /translation="YPYDVPDYAGYPYDVPDYAGSYPYDVPDYA"
     primer_bind     complement(1748..1801)
                     /label=Hg-002-Mfront
                     /note="color: black; sequence: 
                     CATATGGATAGGATCCTGCATAGTCCGGGACGTCATAGGGATAGCCAGCGTAAT"
     primer_bind     1786..1837
                     /label=Hg-012-CSTN2-F1R
                     /note="color: black; sequence: 
                     GGATCCTATCCATATGACGTTCCAGATTACGCTCTGCCTGGGCGGCTGTGCT"
     misc_feature    1819..1875
                     /label=signal peptide
                     /note="color: #a6acb3"
     primer_bind     2744..2763
                     /label=F1
                     /note="color: black; sequence: GTGACCGGGAGACCTACTCT"
     primer_bind     2842..2861
                     /label=Hg-014-CSTN2-F2/F4
                     /note="color: black; sequence: TGGACTGCAGGACTGCTGGT"
     primer_bind     complement(2842..2861)
                     /label=Hg-013-CSTN2-R1/R3
                     /note="color: black; sequence: ACCAGCAGTCCTGCAGTCCA"
     primer_bind     complement(3035..3054)
                     /label=R1
                     /note="color: black; sequence: GTGATGCCGGTTCATTTCGG"
     primer_bind     complement(4664..4698)
                     /label=Hg-015-CSTN2-R2
                     /note="color: black; sequence: 
                     GCACCTGAGGAGTGCCTAGTAGGGGAGGGTGGAGT"
     misc_feature    4684..4698
                     /label=recombination fragment rear
                     /note="color: #00ff00; direction: RIGHT"
     primer_bind     complement(4723..4742)
                     /label=CAG-R
                     /note="CAG-R"
                     /note="color: black; sequence: CAGGGCATTGGCCACACCAG"
     polyA_site      4828..4883
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal"
                     /note="color: #a6acb3"
     primer_bind     complement(5242..5258)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
                     /note="color: #a020f0; direction: LEFT"
     protein_bind    5266..5282
                     /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)."
                     /note="color: #31849b"
     promoter        complement(5290..5320)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
                     /note="This reverse directional feature has 3 segments:
                        1: 5290 .. 5296 / #ffffff / -10
                        2: 5297 .. 5314 / #ffffff
                        3: 5315 .. 5320 / #ffffff / -35"
     protein_bind    5335..5356
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
                     /note="color: #31849b"
     promoter        5415..5610
                     /label=SV40 promoter
                     /note="SV40 early promoter"
                     /note="color: #ffffff; direction: RIGHT"
     rep_origin      5461..5596
                     /label=SV40 ori
                     /note="SV40 origin of replication"
                     /note="color: #ffff00"
     polyA_signal    5616..5750
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
                     /note="color: #a6acb3"
     rep_origin      complement(5989..6577)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
                     /note="color: #ffff00"
     CDS             complement(6748..7608)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /note="This feature has 2 segments:
                        1: 6748 .. 7539 / #ccffcc
                        2: 7540 .. 7608 / #ccffcc / signal sequence
                     Cleavage site after base 7539"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     promoter        complement(7609..7713)
                     /gene="bla"
                     /label=AmpR promoter
                     /note="color: #ffffff; direction: LEFT"
ORIGIN
        1 gtcgacattg attattgact agttattaat agtaatcaat tacggggtca ttagttcata
       61 gcccatatat ggagttccgc gttacataac ttacggtaaa tggcccgcct ggctgaccgc
      121 ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt tcccatagta acgccaatag
      181 ggactttcca ttgacgtcaa tgggtggact atttacggta aactgcccac ttggcagtac
      241 atcaagtgta tcatatgcca agtacgcccc ctattgacgt caatgacggt aaatggcccg
      301 cctggcatta tgcccagtac atgaccttat gggactttcc tacttggcag tacatctacg
      361 tattagtcat cgctattacc atgggtcgag gtgagcccca cgttctgctt cactctcccc
      421 atctcccccc cctccccacc cccaattttg tatttattta ttttttaatt attttgtgca
      481 gcgatggggg cggggggggg gggggcgcgc gccaggcggg gcggggcggg gcgaggggcg
      541 gggcggggcg aggcggagag gtgcggcggc agccaatcag agcggcgcgc tccgaaagtt
      601 tccttttatg gcgaggcggc ggcggcggcg gccctataaa aagcgaagcg cgcggcgggc
      661 gggagtcgct gcgttgcctt cgccccgtgc cccgctccgc gccgcctcgc gccgcccgcc
      721 ccggctctga ctgaccgcgt tactcccaca ggtgagcggg cgggacggcc cttctcctcc
      781 gggctgtaat tagcgcttgg tttaatgacg gctcgtttct tttctgtggc tgcgtgaaag
      841 ccttaaaggg ctccgggagg gccctttgtg cgggggggag cggctcgggg ggtgcgtgcg
      901 tgtgtgtgtg cgtggggagc gccgcgtgcg gcccgcgctg cccggcggct gtgagcgctg
      961 cgggcgcggc gcggggcttt gtgcgctccg cgtgtgcgcg aggggagcgc ggccgggggc
     1021 ggtgccccgc ggtgcggggg ggctgcgagg ggaacaaagg ctgcgtgcgg ggtgtgtgcg
     1081 tgggggggtg agcagggggt gtgggcgcgg cggtcgggct gtaacccccc cctgcacccc
     1141 cctccccgag ttgctgagca cggcccggct tcgggtgcgg ggctccgtgc ggggcgtggc
     1201 gcggggctcg ccgtgccggg cggggggtgg cggcaggtgg gggtgccggg cggggcgggg
     1261 ccgcctcggg ccggggaggg ctcgggggag gggcgcggcg gccccggagc gccggcggct
     1321 gtcgaggcgc ggcgagccgc agccattgcc ttttatggta atcgtgcgag agggcgcagg
     1381 gacttccttt gtcccaaatc tggcggagcc gaaatctggg aggcgccgcc gcaccccctc
     1441 tagcgggcgc gggcgaagcg gtgcggcgcc ggcaggaagg aaatgggcgg ggagggcctt
     1501 cgtgcgtcgc cgcgccgccg tccccttctc catctccagc ctcggggctg ccgcaggggg
     1561 acggctgcct tcggggggga cggggcaggg cggggttcgg cttctggcgt gtgaccggcg
     1621 gctctagagc ctctgctaac catgttcatg ccttcttctt tttcctacag ctcctgggca
     1681 acgtgctggt tattgtgctg tctcatcatt ttggcaaagg ccaccatgta cccatacgat
     1741 gttccagatt acgctggcta tccctatgac gtcccggact atgcaggatc ctatccatat
     1801 gacgttccag attacgctct gcctgggcgg ctgtgctggg tgccgctcct gctggcgctg
     1861 ggcgtgggga gcggcagcgg cggtggcggg gacagccggc agcgccgcct cctcgcggct
     1921 aaagtcaata agcacaagcc atggatcgag acttcatatc atggagtcat aactgagaac
     1981 aatgacacag tcattttgga cccaccactg gtagccctgg ataaagatgc accggttcct
     2041 tttgcagggg aaatctgtgc gttcaagatc catggccagg agctgccctt tgaggctgtg
     2101 gtgctcaaca agacatcagg agagggccgg ctccgtgcca agagccccat tgactgtgag
     2161 ttgcagaagg agtacacatt catcatccag gcctatgact gtggtgctgg gccccacgag
     2221 acagcctgga aaaagtcaca caaggccgtg gtccatatac aggtgaagga tgtcaacgag
     2281 tttgctccca ccttcaaaga gccagcctac aaggctgttg tgacggaggg caagatctat
     2341 gacagcattc tgcaggtgga ggccattgac gaggactgct ccccacagta cagccagatc
     2401 tgcaactatg aaatcgtcac cacagatgtg ccttttgcca tcgacagaaa tggcaacatc
     2461 aggaacactg agaagctgag ctatgacaaa caacaccagt atgagatcct ggtgaccgcc
     2521 tacgactgtg gacagaagcc cgctgctcag gacaccctgg tgcaggtgga tgtgaagcca
     2581 gtttgcaagc ctggctggca agactggacc aagaggattg agtaccagcc tggctccggg
     2641 agcatgcccc tgttccccag catccacctg gagacgtgcg atggagccgt gtcttccctc
     2701 cagatcgtca cagagctgca gactaattac attgggaagg gttgtgaccg ggagacctac
     2761 tctgagaaat cccttcaaaa gttatgtgga gcctcctctg gcatcattga cctcttgcca
     2821 tcccctagcg ctgccaccaa ctggactgca ggactgctgg tggacagcag tgagatgatc
     2881 ttcaagtttg acggcaggca gggtgccaaa gtccccgatg ggattgtgcc caagaacctg
     2941 accgatcagt tcaccatcac catgtggatg aaacacggcc ccagccctgg tgtgagagcc
     3001 gagaaggaaa ccatcctctg caactcagac aaaaccgaaa tgaaccggca tcactatgcc
     3061 ctgtatgtgc acaactgccg cctcgtcttt ctcttgcgga aggacttcga ccaggctgac
     3121 acctttcgcc ccgcggagtt ccactggaag ctggatcaga tttgtgacaa agagtggcac
     3181 tactatgtca tcaatgtgga gtttcctgtg gtaaccttat acatggatgg agcaacatat
     3241 gaaccatacc tggtgaccaa cgactggccc attcatccat ctcacatagc catgcaactc
     3301 acagtcggcg cttgttggca aggaggagaa gtcaccaaac cacagtttgc tcagttcttt
     3361 catggaagcc tggccagtct caccatccgc cctggcaaaa tggaaagcca gaaggtgatc
     3421 tcctgcctgc aggcctgcaa ggaagggctg gacattaatt ccttggaaag ccttggccaa
     3481 ggaataaagt atcacttcaa cccctcgcag tccatcctgg tgatggaagg tgacgacatt
     3541 gggaacatta accgtgctct ccagaaagtc tcctacatca actccaggca gttcccaacg
     3601 gcgggtgtgc ggcgcctcaa agtatcctcc aaagtccagt gctttgggga agacgtatgc
     3661 atcagtatcc ctgaggtaga tgcctatgtg atggtcctcc aggccatcga gccccggatc
     3721 accctccggg gcacagacca cttctggaga cctgctgccc agtttgaaag tgccagggga
     3781 gtgaccctct tccctgatat caagattgtg agcaccttcg ccaaaaccga agcccccggg
     3841 gacgtgaaaa ccacagaccc caaatcagaa gtcttagagg aaatgcttca taacttagat
     3901 ttctgtgaca ttttggtgat cggaggggac ttggacccaa ggcaggagtg cttggagctc
     3961 aaccacagtg agctccacca acgacacctg gatgccacta attctactgc aggctactcc
     4021 atctacggtg tgggctccat gagccgctat gagcaggtgc tacatcacat ccgctaccgc
     4081 aactggcgtc cggcttccct tgaggcccgg cgtttccgga ttaagtgctc agaactcaat
     4141 gggcgctaca ctagcaatga gttcaacttg gaggtcagca tccttcatga agaccaagtc
     4201 tcagataagg agcatgtcaa tcatctgatt gtgcagcctc ccttcctcca gtctgtccat
     4261 catcctgagt cccggagtag catccagcac agttcagtgg tcccaagcat tgccacagtg
     4321 gtcatcatca tctccgtgtg catgcttgtg tttgtcgtgg ccatgggtgt gtaccgggtc
     4381 cggatcgccc accagcactt catccaggag actgaggctg ccaaggaatc tgagatggac
     4441 tgggacgatt ctgcgctgac tatcacagtc aaccccatgg agaaacatga aggaccaggg
     4501 catggggaag atgagactga gggagaagag gaggaagaag ccgaggaaga aatgagctcc
     4561 agcagtggct ctgacgacag cgaagaggag gaggaggagg aagggatggg cagaggcaga
     4621 catgggcaga atggagccag gcaagcccag ctggagtggg atgactccac cctcccctac
     4681 taggcactcc tcaggtgcag gctgcctatc agaaggtggt ggctggtgtg gccaatgccc
     4741 tggctcacaa ataccactga gatctttttc cctctgccaa aaattatggg gacatcatga
     4801 agccccttga gcatctgact tctggctaat aaaggaaatt tattttcatt gcaatagtgt
     4861 gttggaattt tttgtgtctc tcactcggaa ggacatatgg gagggcaaat catttaaaac
     4921 atcagaatga gtatttggtt tagagtttgg caacatatgc catatgctgg ctgccatgaa
     4981 caaaggtggc tataaagagg tcatcagtat atgaaacagc cccctgctgt ccattcctta
     5041 ttccatagaa aagccttgac ttgaggttag atttttttta tattttgttt tgtgttattt
     5101 ttttctttaa catccctaaa attttcctta catgttttac tagccagatt tttcctcctc
     5161 tcctgactac tcccagtcat agctgtccct cttctcttat gaagatccct cgacctgcag
     5221 cccaagcttg gcgtaatcat ggtcatagct gtttcctgtg tgaaattgtt atccgctcac
     5281 aattccacac aacatacgag ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt
     5341 gagctaactc acattaattg cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc
     5401 gtgccagcgg atccgcatct caattagtca gcaaccatag tcccgcccct aactccgccc
     5461 atcccgcccc taactccgcc cagttccgcc cattctccgc cccatggctg actaattttt
     5521 tttatttatg cagaggccga ggccgcctcg gcctctgagc tattccagaa gtagtgagga
     5581 ggcttttttg gaggcctagg cttttgcaaa aagctaactt gtttattgca gcttataatg
     5641 gttacaaata aagcaatagc atcacaaatt tcacaaataa agcatttttt tcactgcatt
     5701 ctagttgtgg tttgtccaaa ctcatcaatg tatcttatca tgtctggatc cgctgcatta
     5761 atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc
     5821 gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa
     5881 ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa
     5941 aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct
     6001 ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac
     6061 aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc
     6121 gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc
     6181 tcaatgctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg
     6241 tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga
     6301 gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta acaggattag
     6361 cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta
     6421 cactagaagg acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag
     6481 agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg
     6541 caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac
     6601 ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc
     6661 aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag
     6721 tatatatgag taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc
     6781 agcgatctgt ctatttcgtt catccatagt tgcctgactc cccgtcgtgt agataactac
     6841 gatacgggag ggcttaccat ctggccccag tgctgcaatg ataccgcgag acccacgctc
     6901 accggctcca gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg
     6961 tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag
     7021 tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc
     7081 acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac
     7141 atgatccccc atgttgtgca aaaaagcggt tagctccttc ggtcctccga tcgttgtcag
     7201 aagtaagttg gccgcagtgt tatcactcat ggttatggca gcactgcata attctcttac
     7261 tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg
     7321 agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaatacggg ataataccgc
     7381 gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact
     7441 ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg cacccaactg
     7501 atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa
     7561 tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcatac tcttcctttt
     7621 tcaatattat tgaagcattt atcagggtta ttgtctcatg agcggataca tatttgaatg
     7681 tatttagaaa aataaacaaa taggggttcc gcgcacattt ccccgaaaag tgccacctg
//
