LOCUS       sequence-437015-        5600 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  A plasmid encoding the hydrogel-forming protein PXP with both the 
            T40A and Q54A coil mutations.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5600)
  AUTHORS   Bennett JI, Boit MO, Gregorio NE, Zhang F, Kibler RD, Hoye JW, Prado
            O, Rapp PB, Murry CE, Stevens KR, DeForest CA
  TITLE     Genetically Encoded XTEN-based Hydrogels with Tunable 
            Viscoelasticity and Biodegradability for Injectable Cell Therapies.
  JOURNAL   Adv Sci (Weinh). 2024 Mar 13:e2301708. doi: 10.1002/advs.202301708.
  PUBMED    38477407
REFERENCE   2  (bases 1 to 5600)
  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: "Adv Sci 
            (Weinh). 2024 Mar 13:e2301708. doi: 10.1002/advs.202301708."
COMMENT     Sequence Label: T40A+Q54A
FEATURES             Location/Qualifiers
     source          1..5600
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(10..24,25..30,31..47,48..54)
                     /label=T5 promoter
                     /note="bacteriophage T5 promoter for E. coli RNA 
                     polymerase, with embedded lac operator"
     protein_bind    30..46
                     /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)."
     protein_bind    62..78
                     /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             97..108
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             127..144
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1018..1035
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     terminator      1057..1151
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             1195..1854
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(1261..1280)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     terminator      1919..2005
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     CDS             complement(2098..3180)
                     /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     3142..3161
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(3181..3258)
                     /gene="lacI (mutant)"
                     /label=lacIq promoter
                     /note="In the lacIq allele, a single base change in the 
                     promoter boosts expression of the lacI gene about 10-fold."
     primer_bind     complement(3342..3364)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    3450..3590
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     3505..3524
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(3605..3622)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3776..4364)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3856..3875)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4535..5326,5327..5395))
                     /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     5158..5177
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5396..5500)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5568..5586
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 gtcgagaaat cataaaaaat ttatttgctt tgtgagcgga taacaattat aatagattca
       61 attgtgagcg gataacaatt tcacacagaa ttcattaaag aggagaaatt aactatgaga
      121 ggatcgcatc accatcacca tcacggatcc ggtcgtggtg actctggctc aggttcgggt
      181 agtgggagcg gctctggcag cggtgcgccg caaatgctgc gtgaactgca ggaagccaat
      241 gccgcgcttc aggatgtgcg ggaattgctt cgtcaagcgg tcaaggagat aacgttcttg
      301 aagaacaccg tcatggagtc ggatgcgtct ggctccggaa gcggaagtgg ttctggtagc
      361 gtcgacggct ccccggctgg cagcccgacc agcactgaag agggcacgag cgagtcggcg
      421 accccggagt ctggtccggg cacctccacc gaaccgtctg agggcagcgc accgggtagc
      481 ccggccggta gccctaccag caccgaagag ggtaccagca cggaaccgag cgaaggctcg
      541 gcaccgggta cgagcaccga gccgtccgag ggttccgcgc caggtaccag cgagagcgca
      601 acgccggagt ccggtccggg cagcgaacca gcgaccagcg gcagcgaaac gccgggttca
      661 gagccggcga cgagcggtag cgagactccg ggcagcccgg ctggtagccc gacgtccacc
      721 gaagaaggca ccagcgaaag cgccaccccg gagagcggtc ctggtacgtc taccgagcca
      781 tctgagggta gcgcgccgct cgagtcaggt tcgggtagtg ggagcggctc tggcagcggt
      841 gcgccgcaaa tgctgcgtga actgcaggaa gccaatgccg cgcttcagga tgtgcgggaa
      901 ttgcttcgtc aagcggtcaa ggagataacg ttcttgaaga acaccgtcat ggagtcggat
      961 gcgtccggat ctggctccgg aagcggaagt ggttctggta gcggtcgtgg tgactctcat
     1021 caccatcacc atcacaagct taattagctg agcttggact cctgttgata gatccagtaa
     1081 tgacctcaga actccatctg gatttgttca gaacgctcgg ttgccgccgg gcgtttttta
     1141 ttggtgagaa tccaagctag cttggcgaga ttttcaggag ctaaggaagc taaaatggag
     1201 aaaaaaatca ctggatatac caccgttgat atatcccaat ggcatcgtaa agaacatttt
     1261 gaggcatttc agtcagttgc tcaatgtacc tataaccaga ccgttcagct ggatattacg
     1321 gcctttttaa agaccgtaaa gaaaaataag cacaagtttt atccggcctt tattcacatt
     1381 cttgcccgcc tgatgaatgc tcatccggaa tttcgtatgg caatgaaaga cggtgagctg
     1441 gtgatatggg atagtgttca cccttgttac accgttttcc atgagcaaac tgaaacgttt
     1501 tcatcgctct ggagtgaata ccacgacgat ttccggcagt ttctacacat atattcgcaa
     1561 gatgtggcgt gttacggtga aaacctggcc tatttcccta aagggtttat tgagaatatg
     1621 tttttcgtct cagccaatcc ctgggtgagt ttcaccagtt ttgatttaaa cgtggccaat
     1681 atggacaact tcttcgcccc cgttttcacc atgggcaaat attatacgca aggcgacaag
     1741 gtgctgatgc cgctggcgat tcaggttcat catgccgttt gtgatggctt ccatgtcggc
     1801 agaatgctta atgaattaca acagtactgc gatgagtggc agggcggggc gtaatttttt
     1861 taaggcagtt attggtgccc ttaaacgcct ggggtaatga ctctctagct tgaggcatca
     1921 aataaaacga aaggctcagt cgaaagactg ggcctttcgt tttatctgtt gtttgtcggt
     1981 gaacgctctc ctgagtagga caaatccgcc ctctagatta cgtgcagtcg atgataagct
     2041 gtcaaacatg agaattgtgc ctaatgagtg agctaactta cattaattgc gttgcgctca
     2101 ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc attaatgaat cggccaacgc
     2161 gcggggagag gcggtttgcg tattgggcgc cagggtggtt tttcttttca ccagtgagac
     2221 gggcaacagc tgattgccct tcaccgcctg gccctgagag agttgcagca agcggtccac
     2281 gctggtttgc cccagcaggc gaaaatcctg tttgatggtg gttaacggcg ggatataaca
     2341 tgagctgtct tcggtatcgt cgtatcccac taccgagata tccgcaccaa cgcgcagccc
     2401 ggactcggta atggcgcgca ttgcgcccag cgccatctga tcgttggcaa ccagcatcgc
     2461 agtgggaacg atgccctcat tcagcatttg catggtttgt tgaaaaccgg acatggcact
     2521 ccagtcgcct tcccgttccg ctatcggctg aatttgattg cgagtgagat atttatgcca
     2581 gccagccaga cgcagacgcg ccgagacaga acttaatggg cccgctaaca gcgcgatttg
     2641 ctggtgaccc aatgcgacca gatgctccac gcccagtcgc gtaccgtctt catgggagaa
     2701 aataatactg ttgatgggtg tctggtcaga gacatcaaga aataacgccg gaacattagt
     2761 gcaggcagct tccacagcaa tggcatcctg gtcatccagc ggatagttaa tgatcagccc
     2821 actgacgcgt tgcgcgagaa gattgtgcac cgccgcttta caggcttcga cgccgcttcg
     2881 ttctaccatc gacaccacca cgctggcacc cagttgatcg gcgcgagatt taatcgccgc
     2941 gacaatttgc gacggcgcgt gcagggccag actggaggtg gcaacgccaa tcagcaacga
     3001 ctgtttgccc gccagttgtt gtgccacgcg gttgggaatg taattcagct ccgccatcgc
     3061 cgcttccact ttttcccgcg ttttcgcaga aacgtggctg gcctggttca ccacgcggga
     3121 aacggtctga taagagacac cggcatactc tgcgacatcg tataacgtta ctggtttcac
     3181 attcaccacc ctgaattgac tctcttccgg gcgctatcat gccataccgc gaaaggtttt
     3241 gcaccattcg atggtgtcgg aatttcgggc agcgttgggt cctggccacg ggtgcgcatg
     3301 atctagagct gcctcgcgcg tttcggtgat gacggtgaaa acctctgaca catgcagctc
     3361 ccggagacgg tcacagcttg tctgtaagcg gatgccggga gcagacaagc ccgtcagggc
     3421 gcgtcagcgg gtgttggcgg gtgtcggggc gcagccatga cccagtcacg tagcgatagc
     3481 ggagtgtata ctggcttaac tatgcggcat cagagcagat tgtactgaga gtgcaccata
     3541 tgcggtgtga aataccgcac agatgcgtaa ggagaaaata ccgcatcagg cgctcttccg
     3601 cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc
     3661 actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga aagaacatgt
     3721 gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc
     3781 ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa
     3841 acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc
     3901 ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg
     3961 cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc
     4021 tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc
     4081 gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca
     4141 ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact
     4201 acggctacac tagaaggaca gtatttggta tctgcgctct gctgaagcca gttaccttcg
     4261 gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt
     4321 ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct
     4381 tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga
     4441 gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa
     4501 tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac
     4561 ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga
     4621 taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcgagacc
     4681 cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca
     4741 gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta
     4801 gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct acaggcatcg
     4861 tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc
     4921 gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg
     4981 ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt
     5041 ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt
     5101 cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca atacgggata
     5161 ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc
     5221 gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac
     5281 ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa
     5341 ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct
     5401 tcctttttca atattattga agcatttatc agggttattg tctcatgagc ggatacatat
     5461 ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc
     5521 cacctgacgt ctaagaaacc attattatca tgacattaac ctataaaaat aggcgtatca
     5581 cgaggccctt tcgtcttcac
//
