LOCUS       sequence-249010-        6831 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  mRNA synthesis.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6831)
  AUTHORS   Horb ME, Thomsen GH
  TITLE     A vegetally localized T-box transcription factor in Xenopus eggs 
            specifies mesoderm and endoderm and is essential for embryonic 
            mesoderm formation.
  JOURNAL   Development. 1997 May;124(9):1689-98.
  PUBMED    9165117
REFERENCE   2  (bases 1 to 6831)
  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: 
            "Development."; date: "1997-05"; volume: "124(9)"; pages: "1689-98"
COMMENT     Sequence Label: pCS2-BRAT
FEATURES             Location/Qualifiers
     source          1..6831
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        35..53
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     35..52
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     RBS             1707..1715
                     /label=Shine-Dalgarno sequence
                     /note="full consensus sequence for ribosome-binding sites 
                     upstream of start codons in E. coli; complementary to a 
                     region in the 3' end of the 16S rRNA (Chen et al., 1994)"
     polyA_signal    2883..3017
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2907..2926)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2961..2980
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(3130..3150)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(3130..3148)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(3169..3185)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3169..3185)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3182..3204)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3193..3209
                     /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(3217..3223,3224..3241,3242..3247))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3262..3283
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3400..3417)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3571..4159)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3651..3670)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4330..5121,5122..5190))
                     /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     4953..4972
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5191..5295)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(5321..5776)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5458..5479)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5670..5689
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        5847..6831
                     /label=CMV IE94 promoter
                     /note="enhancer/promoter region of simian cytomegalovirus 
                     major immediate early transcription unit IE94"
ORIGIN
        1 cgccattctg cctggggacg tcggagcaag cttgatttag gtgacactat agaatacaag
       61 ctacttgttc tttttgcagg atcccatcga ttcgaattca aggtcgacac tagtggatcc
      121 aaagaattcg gcacgagcac ctgcagcaga gtttttatat tgagcattca cctgcagggg
      181 cccatgtgat agcctatagc tacagtgctg gaatgagaaa ctgctgtcgg gaatgcggcc
      241 tctctgcggg gcatctggaa ccagaggctt cttctaactg tgcatcagat gtaaagtctt
      301 ccccagacat ggacagtgtc tccagccaag actccctcta cctgcccaac actgttggtg
      361 cctccctgga agaccaagat ctatggtccc agttccacca ggaggggaca gagatgatca
      421 tcaccaaatc tggaaggagg atgtttcctc agtgtaagat ccgtctcttc ggtcttcatc
      481 cttatgccaa gtacatgctg ctggtggact ttgttcctct ggacaacttt aggtataagt
      541 ggaataagaa ccagtgggaa gcagcaggta aagcagaacc tcacccaccc tgcaggacgt
      601 atgtccaccc agattcacct gctcctggtg cccactggat gaaggatccg atctgctttc
      661 agaagctcaa actcaccaac aacacattgg atcaacaagg ccatattatc ttgcattcaa
      721 tgcatcgcta caagcccagg ttccatgtag ttcagtctga tgacatgtac aattctccat
      781 ggggattggt acaagtgttt agcttcccag agacagagtt tacttcagtg actgcctacc
      841 agaatgaaaa gattactaaa ctgaaaatta atcacaaccc atttgctaaa ggattccggg
      901 agcaggaaag gagtcacaag agggatgatg ttttaaagat tctacaacaa agtcccagta
      961 aaaggcagaa gaggaagaag tgggaggaca gtcctgaggc tgatatttca gatttcccca
     1021 aggctatatg tgtgaaggag gaatccatta tggacccagc aggagtttat cagaactggg
     1081 tttcagatca cgaggctaac caaggcttga caccccactc ccctgagtct gagggtgcca
     1141 atcaggagca gcaagtcccc acatcttcct ctaacttcta caacaagagc cattatcgaa
     1201 ggagttccca acatctctcc tcgccatttg aattgggaga gccctctagc agacgtctta
     1261 cccctgacat tgctacagtg ccggattccg atccagattc tttagcagtg tttcatgtca
     1321 ttccaacaca gaattctgct ccagaacgca catgttctat gaacttttcc atggaagctc
     1381 caatgaaaca gcccctccgg ggtgccatgt acagccctta tggagcagat cagtggttgg
     1441 ttccagctca aggtcaatat cgacctgtgg gctatactgc atacccaaca gacttaagca
     1501 cacaaggagc agtagctcac ccacatagtg cgatgtcaga ctggagccaa tactctctct
     1561 tcccctacag ctgttggtaa atgggttaag ggaaatgtgt atccacagtc cttacccatc
     1621 catgccaata cctcagtcct gatttcttgt tggggggggg ggggagcggg ctgtgtggtt
     1681 ttgcatctct gaggcaaaac ctcaggacct ccttaaaaac cctgcattgg gaggttgcat
     1741 ggagaggttt gattacaata tttgttggac taaaaggtgt cactttatca gcacaaacag
     1801 taggttgccc ctgtgcttgt gatcagggga atctctacct gtacttgaac taaatgtatt
     1861 tattttaaag attagcaagt aaatgtgaga aaccgtgggg gggtgcgtta aagcagatct
     1921 ccagaagaga acccgggaaa gcagttatgg atttatagaa caaagtaatg gggccatgaa
     1981 cagaagcagt tttctcttga ctttaagtgg tggtacagcc atctaactct ccattttctt
     2041 aatgtcccca attattgcct ttcactcaat tatactatgg tctgttttgt tttttgtttt
     2101 tttaaactat aattttgatt acagttttgc accttacatt ttccagggtg gtggtttgca
     2161 gcactaatct gtgacagtgt gtacagcagg tgtttgtgct tttgtgcact gctcgggaaa
     2221 tgtgtgtgta tttgggggga gtgtgtgttc actaaagatt gtactgctga ctatgagact
     2281 aacgctgtgc aatattcaca gggataggca aaatttggtg ttccatctaa agcaaagcta
     2341 cagcttccag tattgcctct agatgagagc actacttctg caatagcagg gtcaccaaag
     2401 gctgcctaac tctgcatatt ttcacattag cataacctct ggcaatgatt gggacaattg
     2461 actccaaaag tggggtaaac atgtaacaaa tattaatcct tgcattctgt gattagttaa
     2521 ataaacatgg gatggtttgt ttgctctttg ttctacagat tcagtagcag ctcagcagtt
     2581 aaagggtttg tgtaattgct tgtaaagcta ggttttgtgg gagggctgca ggcagaacag
     2641 tatgtattct gtatagaaca catctgtaaa tagattttat ttaagaattg tttttgacta
     2701 tgaaatgtgt tttaatgtga cttttattta atggcatgaa ctgaggaagg aaacaatgaa
     2761 gtgtcatttg ttttgtctcc atctggttat acaccttcaa taaatgtttt ccttatttca
     2821 aaaaaaaaaa aaaaaaactc gacctctcga gcctctagaa ctatagtgag tcgtattacg
     2881 tagatccaga catgataaga tacattgatg agtttggaca aaccacaact agaatgcagt
     2941 gaaaaaaatg ctttatttgt gaaatttgtg atgctattgc tttatttgta accattataa
     3001 gctgcaataa acaagttaac aacaacaatt gcattcattt tatgtttcag gttcaggggg
     3061 aggtgtggga ggttttttaa ttcgcggccg cggcgccaat gcattgggcc cggtacccag
     3121 cttttgttcc ctttagtgag ggttaattgc gcgcttggcg taatcatggt catagctgtt
     3181 tcctgtgtga aattgttatc cgctcacaat tccacacaac atacgagccg ggagcataaa
     3241 gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca ttaattgcgt tgcgctcact
     3301 gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc
     3361 ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg
     3421 ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa tacggttatc
     3481 cacagaatca ggggataacg caggaaagaa catgtgagca aaaggccagc aaaaggccag
     3541 gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca
     3601 tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca
     3661 ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg
     3721 atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcatagct cacgctgtag
     3781 gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt
     3841 tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca
     3901 cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg
     3961 cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa gaacagtatt
     4021 tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc
     4081 cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg
     4141 cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg acgctcagtg
     4201 gaacgaaaac tcacgttaag ggattttggt catgagatta tcaaaaagga tcttcaccta
     4261 gatcctttta aattaaaaat gaagttttaa atcaatctaa agtatatatg agtaaacttg
     4321 gtctgacagt taccaatgct taatcagtga ggcacctatc tcagcgatct gtctatttcg
     4381 ttcatccata gttgcctgac tccccgtcgt gtagataact acgatacggg agggcttacc
     4441 atctggcccc agtgctgcaa tgataccgcg agacccacgc tcaccggctc cagatttatc
     4501 agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc
     4561 ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc cagttaatag
     4621 tttgcgcaac gttgttgcca ttgctacagg catcgtggtg tcacgctcgt cgtttggtat
     4681 ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc ccatgttgtg
     4741 caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt
     4801 gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc catccgtaag
     4861 atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt gtatgcggcg
     4921 accgagttgc tcttgcccgg cgtcaatacg ggataatacc gcgccacata gcagaacttt
     4981 aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct
     5041 gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag catcttttac
     5101 tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat
     5161 aagggcgaca cggaaatgtt gaatactcat actcttcctt tttcaatatt attgaagcat
     5221 ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga aaaataaaca
     5281 aataggggtt ccgcgcacat ttccccgaaa agtgccacct aaattgtaag cgttaatatt
     5341 ttgttaaaat tcgcgttaaa tttttgttaa atcagctcat tttttaacca ataggccgaa
     5401 atcggcaaaa tcccttataa atcaaaagaa tagaccgaga tagggttgag tgttgttcca
     5461 gtttggaaca agagtccact attaaagaac gtggactcca acgtcaaagg gcgaaaaacc
     5521 gtctatcagg gcgatggccc actacgtgaa ccatcaccct aatcaagttt tttggggtcg
     5581 aggtgccgta aagcactaaa tcggaaccct aaagggagcc cccgatttag agcttgacgg
     5641 ggaaagccgg cgaacgtggc gagaaaggaa gggaagaaag cgaaaggagc gggcgctagg
     5701 gcgctggcaa gtgtagcggt cacgctgcgc gtaaccacca cacccgccgc gcttaatgcg
     5761 ccgctacagg gcgcgtccca ttcgccattc aggctgcgca actgttggga agggcgatcg
     5821 gtgcgggcct cttcgctatt acgccagtcg accgccaatt caatatggcg tatatggact
     5881 catgccaatt caatatggtg gatctggacc tgtgccaatt caatatggcg tatatggact
     5941 cgtgccaatt caatatggtg gatctggacc ccagccaatt caatatggcg gacttggcac
     6001 catgccaatt caatatggcg gacctggcac tgtgccaact ggggaggggt ctacttggca
     6061 cggtgccaag tttgaggagg ggtcttggcc ctgtgccaag tccgccatat tgaattggca
     6121 tggtgccaat aatggcggcc atattggcta tatgccagga tcaatatata ggcaatatcc
     6181 aatatggccc tatgccaata tggctattgg ccaggttcaa tactatgtat tggccctatg
     6241 ccatatagta ttccatatat gggttttcct attgacgtag atagcccctc ccaatgggcg
     6301 gtcccatata ccatatatgg ggcttcctaa taccgcccat agccactccc ccattgacgt
     6361 caatggtctc tatatatggt ctttcctatt gacgtcatat gggcggtcct attgacgtat
     6421 atggcgcctc ccccattgac gtcaattacg gtaaatggcc cgcctggctc aatgcccatt
     6481 gacgtcaata ggaccaccca ccattgacgt caatgggatg gctcattgcc cattcatatc
     6541 cgttctcacg ccccctattg acgtcaatga cggtaaatgg cccacttggc agtacatcaa
     6601 tatctattaa tagtaacttg gcaagtacat tactattgga agtacgccag ggtacattgg
     6661 cagtactccc attgacgtca atggcggtaa atggcccgcg atggctgcca agtacatccc
     6721 cattgacgtc aatggggagg ggcaatgacg caaatgggcg ttccattgac gtaaatgggc
     6781 ggtaggcgtg cctaatggga ggtctatata agcaatgctc gtttagggaa c
//
