LOCUS       sequence-192979-        5242 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of human SPDEF truncation.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5242)
  AUTHORS   Lo YH, Noah TK, Chen MS, Zou W, Borras E, Vilar E, Shroyer NF
  TITLE     SPDEF Induces Quiescence of Colorectal Cancer Cells by Changing the 
            Transcriptional Targets of beta-catenin.
  JOURNAL   Gastroenterology. 2017 Jul;153(1):205-218.e8. doi: 
            10.1053/j.gastro.2017.03.048. Epub 2017 Apr 5.
  PUBMED    28390865
REFERENCE   2  (bases 1 to 5242)
  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"; doi: 
            "10.1053/j.gastro.2017.03.048"; journalName: "Gastroenterology"; 
            date: "2017-07"; volume: "153"; issue: "1"; pages: "205-218.e8"
COMMENT     Sequence Label: pCMV7.1-Flag-NLS-hSPDEF delta2
FEATURES             Location/Qualifiers
     source          1..5242
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..380
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        381..584
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     535..555
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             614..679
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             701..721
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     polyA_signal    1266..1888
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(1465..1484)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     primer_bind     complement(1912..1932)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        1917..2246
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2097..2232
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(2284..2303)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2285..2303)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2317..2333)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2317..2333)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2330..2352)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2341..2357
                     /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(2365..2371,2372..2389,2390..2395))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2410..2431
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2548..2565)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2719..3307)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2799..2818)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3478..4269,4270..4338))
                     /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     4101..4120
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4339..4443)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      4470..4925
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4557..4576)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4767..4788
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5051..5073
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     5065..5082
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     5066..5082
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
ORIGIN
        1 gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
       61 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca
      121 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga
      181 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc
      241 aagtgtatca tatgccaagt ccgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
      301 ggcattatgc ccagtacatg accttacggg actttcctac ttggcagtac atctacgtat
      361 tagtcatcgc tattaccatg gtgatgcggt tttggcagta caccaatggg cgtggatagc
      421 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt
      481 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaataa ccccgccccg ttgacgcaaa
      541 tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctcgttta gtgaaccgtc
      601 agaattaacc atggactaca aagaccatga cggtgattat aaagatcatg acatcgatta
      661 caaggatgac gatgacaagc ttgcggccgc gaattccgcc cccaagaaaa agagaaaggt
      721 gagcagctgg gcagccaagg cccctggggc cagcagtcgg gaggagccac ctgaggagcc
      781 tgagcagtgc ccggtcattg acagccaagc cccagcgggc agcctggact tggtgcccgg
      841 cgggctgacc ttggaggagc actcgctgga gcaggtgcag tccatggtgg tgggcgaagt
      901 gctcaaggac atcgagacgg cctgcaagct gctcaacatc accgcagatc ccatggactg
      961 gagccccagc aatgtgcaga agtggctcct gtggacagag caccaatacc ggctgccccc
     1021 catgggcaag gccttccagg agctggcggg taaggagctg tgcgccatgt cggaggagca
     1081 gttccgccag cgctcgcccc tgggtgggga tgtgctgcac gcccacctgg acatctggaa
     1141 gtcagcggcc tggatgaaag agcggacttc acctggggcg attcactact gtgcctcgac
     1201 cagtgaggag agctggaccg acagcgaggt ggactcatca tgctccgggt agccctgagg
     1261 atcccgggtg gcatccctgt gacccctccc cagtgcctct cctggccctg gaagttgcca
     1321 ctccagtgcc caccagcctt gtcctaataa aattaagttg catcattttg tctgactagg
     1381 tgtccttcta taatattatg gggtggaggg gggtggtatg gagcaagggg caagttggga
     1441 agacaacctg tagggcctgc ggggtctatt gggaaccaag ctggagtgca gtggcacaat
     1501 cttggctcac tgcaatctcc gcctcctggg ttcaagcgat tctcctgcct cagcctcccg
     1561 agttgttggg attccaggca tgcatgacca ggctcagcta atttttgttt ttttggtaga
     1621 gacggggttt caccatattg gccaggctgg tctccaactc ctaatctcag gtgatctacc
     1681 caccttggcc tcccaaattg ctgggattac aggcgtgaac cactgctccc ttccctgtcc
     1741 ttctgatttt aaaataacta taccagcagg aggacgtcca gacacagcat aggctacctg
     1801 gccatgccca accggtggga catttgagtt gcttgcttgg cactgtcctc tcatgcgttg
     1861 ggtccactca gtagatgcct gttgaattgg gtacgcggcc agcttggctg tggaatgtgt
     1921 gtcagttagg gtgtggaaag tccccaggct ccccagcagg cagaagtatg caaagcatgc
     1981 atctcaatta gtcagcaacc aggtgtggaa agtccccagg ctccccagca ggcagaagta
     2041 tgcaaagcat gcatctcaat tagtcagcaa ccatagtccc gcccctaact ccgcccatcc
     2101 cgcccctaac tccgcccagt tccgcccatt ctccgcccca tggctgacta atttttttta
     2161 tttatgcaga ggccgaggcc gcctcggcct ctgagctatt ccagaagtag tgaggaggct
     2221 tttttggagg cctaggcttt tgcaaaaagc tcctcgagga actgaaaaac cagaaagtta
     2281 attccctata gtgagtcgta ttaaattcgt aatcatgtca tagctgtttc ctgtgtgaaa
     2341 ttgttatccg ctcacaattc cacacaacat acgagccgga agcataaagt gtaaagcctg
     2401 gggtgcctaa tgagtgagct aactcacatt aattgcgttg cgctcactgc ccgctttcca
     2461 gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg ggagaggcgg
     2521 tttgcgtatt gggcgctctt ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg
     2581 gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg
     2641 ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa
     2701 ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg
     2761 acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc
     2821 tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc
     2881 ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc
     2941 ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg
     3001 ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc
     3061 actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga
     3121 gttcttgaag tggtggccta actacggcta cactagaaga acagtatttg gtatctgcgc
     3181 tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac
     3241 caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg
     3301 atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc
     3361 acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa
     3421 ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta
     3481 ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt
     3541 tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat ctggccccag
     3601 tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag caataaacca
     3661 gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct ccatccagtc
     3721 tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt
     3781 tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag
     3841 ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca aaaaagcggt
     3901 tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt tatcactcat
     3961 ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat gcttttctgt
     4021 gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac cgagttgctc
     4081 ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa aagtgctcat
     4141 cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag
     4201 ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt
     4261 ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg
     4321 gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt atcagggtta
     4381 ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa taggggttcc
     4441 gcgcacattt ccccgaaaag tgccacctga cgcgccctgt agcggcgcat taagcgcggc
     4501 gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc agcgccctag cgcccgctcc
     4561 tttcgctttc ttcccttcct ttctcgccac gttcgccggc tttccccgtc aagctctaaa
     4621 tcgggggctc cctttagggt tccgatttag tgctttacgg cacctcgacc ccaaaaaact
     4681 tgattagggt gatggttcac gtagtgggcc atcgccctga tagacggttt ttcgcccttt
     4741 gacgttggag tccacgttct ttaatagtgg actcttgttc caaactggaa caacactcaa
     4801 ccctatctcg gtctattctt ttgatttata agggattttg ccgatttcgg cctattggtt
     4861 aaaaaatgag ctgatttaac aaaaatttaa cgcgaatttt aacaaaatat taacgcttac
     4921 aatttccatt cgccattcag gctgcgcaac tgttgggaag ggcgatcggt gcgggcctct
     4981 tcgctattac gccagctggc gaaaggggga tgtgctgcaa ggcgattaag ttgggtaacg
     5041 ccagggtttt cccagtcacg acgttgtaaa acgacggcca gtgccaagct gatctataca
     5101 ttgaatcaat attggcaatt agccatatta gtcattggtt atatagcata aatcaatatt
     5161 ggctattggc cattgcatac gttgtatcta tatcataata tgtacattta tattggctca
     5221 tgtccaatat gaccgccatg tt
//
