LOCUS       sequence-273044-        5831 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Engineered form of fibroblast growth factor 2 with improved 
            thermostability.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5831)
  AUTHORS   Kuo HH, Gao X, DeKeyser JM, Fetterman KA, Pinheiro EA, Weddle CJ, 
            Fonoudi H, Orman MV, Romero-Tejeda M, Jouni M, Blancard M, Magdy T, 
            Epting CL, George AL Jr, Burridge PW
  TITLE     Negligible-Cost and Weekend-Free Chemically Defined Human iPSC 
            Culture.
  JOURNAL   Stem Cell Reports. 2020 Jan 9. pii: S2213-6711(19)30446-1. doi: 
            10.1016/j.stemcr.2019.12.007.
  PUBMED    31928950
REFERENCE   2  (bases 1 to 5831)
  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: "Stem Cell 
            Reports. 2020 Jan 9. pii: S2213-6711(19)30446-1. doi: 
            10.1016/j.stemcr.2019.12.007."
COMMENT     Sequence Label: pET-28a(+)-FGF2-G3
FEATURES             Location/Qualifiers
     source          1..5831
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     1..20
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1..19
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    20..44
                     /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)."
     RBS             59..81
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             100..117
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             127..144
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             148..180
                     /codon_start=1
                     /product="leader peptide from bacteriophage T7 gene 10"
                     /label=T7 tag (gene 10 leader)
                     /note="promotes efficient translation in E. coli"
                     /translation="MASMTGGQQMG"
     CDS             696..713
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(766..784)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      780..827
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      864..1319
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(951..970)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1161..1182
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(1411..2226)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(1411..2226)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     2134..2153
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2218..2237
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      2348..2936
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2837..2856
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3090..3107
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3122..3261
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3187..3206)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3347..3369
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3363..3554)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(4363..5445)
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=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)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     5407..5426
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5446..5523)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     5729..5748
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 taatacgact cactataggg gaattgtgag cggataacaa ttcccctcta gaaataattt
       61 tgtttaactt taagaaggag atataccatg ggcagcagcc atcatcatca tcatcacagc
      121 agcggcctgg tgccgcgcgg cagccatatg gctagcatga ctggtggaca gcaaatgggt
      181 cgcggatcca tggcagcagg ttcgatcact acattaccgg cactgccgga agatggtggt
      241 tcaggtgcat ttcctcctgg ccacttcaaa gaccctaaac tgctgtactg caagaatggc
      301 ggcttctttc tgcgcattca cccggatggc cgtgttgatg gtactcgcga taaatcagat
      361 ccgttcatca agctgcagct gcaagcggaa gaacgtggcg tggtgagcat taagggcgtt
      421 tgtgcaaacc gttatttagc gatgaaggaa gacggccgcc tgtacgcgat caagaacgtg
      481 accgacgaat gcttcttctt tgaacgcctg gaagaaaaca actacaacac ctaccgcagc
      541 cgcaagtacc cgagctggta tgttgcgtta aagcgtaccg gccagtataa attaggcagc
      601 aaaaccggtc cgggccagaa ggcgattctg tttctgccta tgagcgcgaa gtcatgaatt
      661 cgagctccgt cgacaagctt gcggccgcac tcgagcacca ccaccaccac cactgagatc
      721 cggctgctaa caaagcccga aaggaagctg agttggctgc tgccaccgct gagcaataac
      781 tagcataacc ccttggggcc tctaaacggg tcttgagggg ttttttgctg aaaggaggaa
      841 ctatatccgg attggcgaat gggacgcgcc ctgtagcggc gcattaagcg cggcgggtgt
      901 ggtggttacg cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc
      961 tttcttccct tcctttctcg ccacgttcgc cggctttccc cgtcaagctc taaatcgggg
     1021 gctcccttta gggttccgat ttagtgcttt acggcacctc gaccccaaaa aacttgatta
     1081 gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt
     1141 ggagtccacg ttctttaata gtggactctt gttccaaact ggaacaacac tcaaccctat
     1201 ctcggtctat tcttttgatt tataagggat tttgccgatt tcggcctatt ggttaaaaaa
     1261 tgagctgatt taacaaaaat ttaacgcgaa ttttaacaaa atattaacgc ttacaattta
     1321 ggtggcactt ttcggggaaa tgtgcgcgga acccctattt gtttattttt ctaaatacat
     1381 tcaaatatgt atccgctcat gaattaattc ttagaaaaac tcatcgagca tcaaatgaaa
     1441 ctgcaattta ttcatatcag gattatcaat accatatttt tgaaaaagcc gtttctgtaa
     1501 tgaaggagaa aactcaccga ggcagttcca taggatggca agatcctggt atcggtctgc
     1561 gattccgact cgtccaacat caatacaacc tattaatttc ccctcgtcaa aaataaggtt
     1621 atcaagtgag aaatcaccat gagtgacgac tgaatccggt gagaatggca aaagtttatg
     1681 catttctttc cagacttgtt caacaggcca gccattacgc tcgtcatcaa aatcactcgc
     1741 atcaaccaaa ccgttattca ttcgtgattg cgcctgagcg agacgaaata cgcgatcgct
     1801 gttaaaagga caattacaaa caggaatcga atgcaaccgg cgcaggaaca ctgccagcgc
     1861 atcaacaata ttttcacctg aatcaggata ttcttctaat acctggaatg ctgttttccc
     1921 ggggatcgca gtggtgagta accatgcatc atcaggagta cggataaaat gcttgatggt
     1981 cggaagaggc ataaattccg tcagccagtt tagtctgacc atctcatctg taacatcatt
     2041 ggcaacgcta cctttgccat gtttcagaaa caactctggc gcatcgggct tcccatacaa
     2101 tcgatagatt gtcgcacctg attgcccgac attatcgcga gcccatttat acccatataa
     2161 atcagcatcc atgttggaat ttaatcgcgg cctagagcaa gacgtttccc gttgaatatg
     2221 gctcataaca ccccttgtat tactgtttat gtaagcagac agttttattg ttcatgacca
     2281 aaatccctta acgtgagttt tcgttccact gagcgtcaga ccccgtagaa aagatcaaag
     2341 gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac
     2401 cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt ccgaaggtaa
     2461 ctggcttcag cagagcgcag ataccaaata ctgtccttct agtgtagccg tagttaggcc
     2521 accacttcaa gaactctgta gcaccgccta catacctcgc tctgctaatc ctgttaccag
     2581 tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac
     2641 cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc agcttggagc
     2701 gaacgaccta caccgaactg agatacctac agcgtgagct atgagaaagc gccacgcttc
     2761 ccgaagggag aaaggcggac aggtatccgg taagcggcag ggtcggaaca ggagagcgca
     2821 cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg tttcgccacc
     2881 tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg
     2941 ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct cacatgttct
     3001 ttcctgcgtt atcccctgat tctgtggata accgtattac cgcctttgag tgagctgata
     3061 ccgctcgccg cagccgaacg accgagcgca gcgagtcagt gagcgaggaa gcggaagagc
     3121 gcctgatgcg gtattttctc cttacgcatc tgtgcggtat ttcacaccgc aatggtgcac
     3181 tctcagtaca atctgctctg atgccgcata gttaagccag tatacactcc gctatcgcta
     3241 cgtgactggg tcatggctgc gccccgacac ccgccaacac ccgctgacgc gccctgacgg
     3301 gcttgtctgc tcccggcatc cgcttacaga caagctgtga ccgtctccgg gagctgcatg
     3361 tgtcagaggt tttcaccgtc atcaccgaaa cgcgcgaggc agctgcggta aagctcatca
     3421 gcgtggtcgt gaagcgattc acagatgtct gcctgttcat ccgcgtccag ctcgttgagt
     3481 ttctccagaa gcgttaatgt ctggcttctg ataaagcggg ccatgttaag ggcggttttt
     3541 tcctgtttgg tcactgatgc ctccgtgtaa gggggatttc tgttcatggg ggtaatgata
     3601 ccgatgaaac gagagaggat gctcacgata cgggttactg atgatgaaca tgcccggtta
     3661 ctggaacgtt gtgagggtaa acaactggcg gtatggatgc ggcgggacca gagaaaaatc
     3721 actcagggtc aatgccagcg cttcgttaat acagatgtag gtgttccaca gggtagccag
     3781 cagcatcctg cgatgcagat ccggaacata atggtgcagg gcgctgactt ccgcgtttcc
     3841 agactttacg aaacacggaa accgaagacc attcatgttg ttgctcaggt cgcagacgtt
     3901 ttgcagcagc agtcgcttca cgttcgctcg cgtatcggtg attcattctg ctaaccagta
     3961 aggcaacccc gccagcctag ccgggtcctc aacgacagga gcacgatcat gcgcacccgt
     4021 ggggccgcca tgccggcgat aatggcctgc ttctcgccga aacgtttggt ggcgggacca
     4081 gtgacgaagg cttgagcgag ggcgtgcaag attccgaata ccgcaagcga caggccgatc
     4141 atcgtcgcgc tccagcgaaa gcggtcctcg ccgaaaatga cccagagcgc tgccggcacc
     4201 tgtcctacga gttgcatgat aaagaagaca gtcataagtg cggcgacgat agtcatgccc
     4261 cgcgcccacc ggaaggagct gactgggttg aaggctctca agggcatcgg tcgagatccc
     4321 ggtgcctaat gagtgagcta acttacatta attgcgttgc gctcactgcc cgctttccag
     4381 tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt
     4441 ttgcgtattg ggcgccaggg tggtttttct tttcaccagt gagacgggca acagctgatt
     4501 gcccttcacc gcctggccct gagagagttg cagcaagcgg tccacgctgg tttgccccag
     4561 caggcgaaaa tcctgtttga tggtggttaa cggcgggata taacatgagc tgtcttcggt
     4621 atcgtcgtat cccactaccg agatatccgc accaacgcgc agcccggact cggtaatggc
     4681 gcgcattgcg cccagcgcca tctgatcgtt ggcaaccagc atcgcagtgg gaacgatgcc
     4741 ctcattcagc atttgcatgg tttgttgaaa accggacatg gcactccagt cgccttcccg
     4801 ttccgctatc ggctgaattt gattgcgagt gagatattta tgccagccag ccagacgcag
     4861 acgcgccgag acagaactta atgggcccgc taacagcgcg atttgctggt gacccaatgc
     4921 gaccagatgc tccacgccca gtcgcgtacc gtcttcatgg gagaaaataa tactgttgat
     4981 gggtgtctgg tcagagacat caagaaataa cgccggaaca ttagtgcagg cagcttccac
     5041 agcaatggca tcctggtcat ccagcggata gttaatgatc agcccactga cgcgttgcgc
     5101 gagaagattg tgcaccgccg ctttacaggc ttcgacgccg cttcgttcta ccatcgacac
     5161 caccacgctg gcacccagtt gatcggcgcg agatttaatc gccgcgacaa tttgcgacgg
     5221 cgcgtgcagg gccagactgg aggtggcaac gccaatcagc aacgactgtt tgcccgccag
     5281 ttgttgtgcc acgcggttgg gaatgtaatt cagctccgcc atcgccgctt ccactttttc
     5341 ccgcgttttc gcagaaacgt ggctggcctg gttcaccacg cgggaaacgg tctgataaga
     5401 gacaccggca tactctgcga catcgtataa cgttactggt ttcacattca ccaccctgaa
     5461 ttgactctct tccgggcgct atcatgccat accgcgaaag gttttgcgcc attcgatggt
     5521 gtccgggatc tcgacgctct cccttatgcg actcctgcat taggaagcag cccagtagta
     5581 ggttgaggcc gttgagcacc gccgccgcaa ggaatggtgc atgcaaggag atggcgccca
     5641 acagtccccc ggccacgggg cctgccacca tacccacgcc gaaacaagcg ctcatgagcc
     5701 cgaagtggcg agcccgatct tccccatcgg tgatgtcggc gatataggcg ccagcaaccg
     5761 cacctgtggc gccggtgatg ccggccacga tgcgtccggc gtagaggatc gagatctcga
     5821 tcccgcgaaa t
//
