LOCUS       sequence-324724-        7119 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  To generate site specific attP-GA integrant cell lines upon 
            co-delivery with BxbI. Contains IRES2-T7-Halo-PDGFR_TM-T2A-Puro. .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7119)
  TITLE     Huang Lab Starport System Plasmids
REFERENCE   2  (bases 1 to 7119)
  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"
COMMENT     Sequence Label: StarShip03
FEATURES             Location/Qualifiers
     source          1..7119
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(29..51)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     151..170
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     364..386
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     378..395
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     379..395
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    1028..1065
                     /label=Bxb1 attB
                     /bound_moiety="Mycobacteriophage Bxb1 integrase"
                     /note="Minimal attB site for the Mycopbacteriophage Bxb1 
                     integrase (Voutev and Mann, 2017)"
     RBS             1333..1344
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             1351..2064
                     /codon_start=1
                     /product="GFP variant that folds robustly even when fused 
                     to poorly folded proteins (Pedelacq et al., 2006)"
                     /label=superfolder GFP
                     /translation="MRKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTISFKDDG
                     TYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIKA
                     NFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSVLSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     terminator      2076..2147
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      2163..2190
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     misc_feature    join(2226..2800,2801..2803,2804..2812)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(2393..2410)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     2620..2639
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             2876..2908
                     /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             2915..3802
                     /codon_start=1
                     /product="modified bacterial dehalogenase that forms 
                     covalent bonds with chloroalkane derivatives"
                     /label=HaloTag(R)
                     /translation="AEIGTGFPFDPHYVEVLGERMHYVDVGPRDGTPVLFLHGNPTSSY
                     VWRNIIPHVAPTHRCIAPDLIGMGKSDKPDLGYFFDDHVRFMDAFIEALGLEEVVLVIH
                     DWGSALGFHWAKRNPERVKGIAFMEFIRPIPTWDEWPEFARETFQAFRTTDVGRKLIID
                     QNVFIEGTLPMGVVRPLTEVEMDHYREPFLNPVDREPLWRFPNELPIAGEPANIVALVE
                     EYMDWLHQSPVPKLLFWGTPGVLIPPAEAARLAKSLPNCKAVDIGPGLNLLQEDNPDLI
                     GSEIARWLSTLEISG"
     CDS             3974..4027
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             4034..4630
                     /codon_start=1
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="TEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIERV
                     TELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLAA
                     QQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETSA
                     PRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     polyA_signal    4639..4773
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4663..4682)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     4717..4736
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(4898..4914)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4898..4914)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4911..4933)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4922..4938
                     /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(4946..4952,4953..4970,4971..4976))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4991..5012
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5129..5146)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5300..5888)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5380..5399)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6059..6850,6851..6919))
                     /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     6682..6701
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6920..7024)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7092..7110
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gagactgtca
       61 cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg
      121 ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta ctgagagtgc
      181 accatatgcg gtgtgaaata ccgcacagat gcgtaaggag aaaataccgc atcaggcgcc
      241 attcgccatt caggctgcgc aactgttggg aagggcgatc ggtgcgggcc tcttcgctat
      301 tacgccagct ggcgaaaggg ggatgtgctg caaggcgatt aagttgggta acgccagggt
      361 tttcccagtc acgacgttgt aaaacgacgg ccagtgaatt gacgcgtatt gggatcactg
      421 cagatcgaat tcgagttggc gcgcctgtca ttctaaatct ctctttcagc ctaaagcttt
      481 ttccccgtat ccccccaggt gtctgcaggc tcaaagagca gcgagaagcg ttcagaggaa
      541 agcgatcccg tgccaccttc cccgtgcccg ggctgtcccc gcacgctgcc ggctcgggga
      601 tgcgggggga gcgccggacc ggagcggagc cccgggcggc tcgctgctgc cccctagcgg
      661 gggagggacg taattacatc cctgggggct ttgggagggg gctgtccccg tgagctccca
      721 ggcgcgcctg tcattctaaa tctctctttc agcctaaagc tttttccccg tatcccccca
      781 ggtgtctgca ggctcaaaga gcagcgagaa gcgttcagag gaaagcgatc ccgtgccacc
      841 ttccccgtgc ccgggctgtc cccgcacgct gccggctcgg ggatgcgggg ggagcgccgg
      901 accggagcgg agccccgggc ggctcgctgc tgccccctag cgggggaggg acgtaattac
      961 atccctgggg gctttggggg ggggctgtcc ccgtgagctc aataaaagag cccactacaa
     1021 acgggccggc ttgtcgacga cggcggactc cgtcgtcagg atcatccggg agctttgcgg
     1081 gttgccattt gtaactccgg atcctaaaat tcctgtcctg ttctctgtct cttctaggtt
     1141 gggggccgtc ccgctcctaa ggcaggaata tgtgagacca gaccgaaagt gaaacgtgat
     1201 ttcatgcgtc attttgaaca ttttgtaaat cttatttaat aatgtgtgcg gcaattcaca
     1261 tttaatttat gaatgttttc ttaacatcgc ggcaactcaa gaaacggcag gttcggatct
     1321 tagctactag agaaagagga gaaatactag atgcgtaaag gcgaagagct gttcactggt
     1381 gtcgtcccta ttctggtgga actggatggt gatgtcaacg gtcataagtt ttccgtgcgt
     1441 ggcgagggtg aaggtgacgc aactaatggt aaactgacgc tgaagttcat ctgtactact
     1501 ggtaaactgc cggttccttg gccgactctg gtaacgacgc tgacttatgg tgttcagtgc
     1561 tttgctcgtt atccggacca tatgaagcag catgacttct tcaagtccgc catgccggaa
     1621 ggctatgtgc aggaacgcac gatttccttt aaggatgacg gcacgtacaa aacgcgtgcg
     1681 gaagtgaaat ttgaaggcga taccctggta aaccgcattg agctgaaagg cattgacttt
     1741 aaagaggacg gcaatatcct gggccataag ctggaataca attttaacag ccacaatgtt
     1801 tacatcaccg ccgataaaca aaaaaatggc attaaagcga attttaaaat tcgccacaac
     1861 gtggaggatg gcagcgtgca gctggctgat cactaccagc aaaacactcc aatcggtgat
     1921 ggtcctgttc tgctgccaga caatcactat ctgagcacgc aaagcgttct gtctaaagat
     1981 ccgaacgaga aacgcgatca tatggttctg ctggagttcg taaccgcagc gggcatcacg
     2041 catggtatgg atgaactgta caaatgacca ggcatcaaat aaaacgaaag gctcagtcga
     2101 aagactgggc ctttcgtttt atctgttgtt tgtcggtgaa cgctctctac tagagtcaca
     2161 ctggctcacc ttcgggtggg cctttctgcg tttataggtc tcatggctag gaattccgcc
     2221 ccccccccct ctccctcccc cccccctaac gttactggcc gaagccgctt ggaataaggc
     2281 cggtgtgcgt ttgtctatat gttattttcc accatattgc cgtcttttgg caatgtgagg
     2341 gcccggaaac ctggccctgt cttcttgacg agcattccta ggggtctttc ccctctcgcc
     2401 aaaggaatgc aaggtctgtt gaatgtcgtg aaggaagcag ttcctctgga agcttcttga
     2461 agacaaacaa cgtctgtagc gaccctttgc aggcagcgga accccccacc tggcgacagg
     2521 tgcctctgcg gccaaaagcc acgtgtataa gatacacctg caaaggcggc acaaccccag
     2581 tgccacgttg tgagttggat agttgtggaa agagtcaaat ggctctcctc aagcgtattc
     2641 aacaaggggc tgaaggatgc ccagaaggta ccccattgta tgggatctga tctggggcct
     2701 cggtgcacat gctttacatg tgtttagtcg aggttaaaaa aacgtctagg ccccccgaac
     2761 cacggggacg tggttttcct ttgaaaaaca cgatgataat atggccacaa ccatggagac
     2821 tgacaccctc ttgctctggg tcctgctgct gtgggttccg ggtagtaccg gtgacatggc
     2881 tagcatgact ggtggacagc aaatgggtgg ttctgcagaa atcggtactg gctttccatt
     2941 cgacccccat tatgtggaag tcctgggcga gcgcatgcac tacgtcgatg ttggtccgcg
     3001 cgatggcacc cctgtgctgt tcctgcacgg taacccgacc tcctcctacg tgtggcgcaa
     3061 catcatcccg catgttgcac cgacccatcg ctgcattgct ccagacctga tcggtatggg
     3121 caaatccgac aaaccagacc tgggttattt cttcgacgac cacgtccgct tcatggatgc
     3181 cttcatcgaa gccctgggtc tggaagaggt cgtcctggtc attcacgact ggggctccgc
     3241 tctgggtttc cactgggcca agcgcaatcc agagcgcgtc aaaggtattg catttatgga
     3301 gttcatccgc cctatcccga cctgggacga atggccagaa tttgcccgcg agactttcca
     3361 ggccttccgc accaccgacg tcggccgcaa gctgatcatc gatcagaacg tttttatcga
     3421 gggtacgctg ccgatgggtg tcgtccgccc gctgactgaa gtcgagatgg accattaccg
     3481 cgagccgttc ctgaatcctg ttgaccgcga gccactgtgg cgcttcccaa acgagctgcc
     3541 aatcgccggt gagccagcga acatcgtcgc gctggtcgaa gaatacatgg actggctgca
     3601 ccagtcccct gtcccgaagc tgctgttctg gggcacccca ggcgttctga tcccaccggc
     3661 cgaagccgct cgcctggcca aaagcctgcc taactgcaag gctgtggaca tcggcccggg
     3721 tctgaatctg ctgcaagagg acaacccgga cctgatcggc agcgagatcg cgcgctggct
     3781 gtcgacgctc gagatttccg gcggttctaa cgccgtgggc caggatacgc aggaggtgat
     3841 cgtcgtgccc cacagcttgc ccttcaaggt cgtggttatt agtgctatcc tggcgttggt
     3901 ggtgctgact attatctccc ttatcatact tatcatgctg tggcaaaaaa aaccccgagg
     3961 atccggaagc ggagagggca gaggaagtct gctaacatgc ggtgacgtcg aggagaatcc
     4021 tggacctggt ggcaccgagt acaagcctac cgtgcgcctg gccactcgcg atgatgtgcc
     4081 ccgcgccgtc cgcactctgg ccgccgcttt cgccgactac cccgctaccc ggcacaccgt
     4141 ggaccccgac cggcacatcg agcgtgtgac agagttgcag gagctgttcc tgacccgcgt
     4201 cgggctggac atcggcaagg tgtgggtagc cgacgacggc gcggccgtgg ccgtgtggac
     4261 tacccccgag agcgttgagg ccggcgccgt gttcgccgag atcggccccc gaatggccga
     4321 gctgagcggc agccgcctgg ccgcccagca gcaaatggag ggcctgcttg ccccccatcg
     4381 tcccaaggag cctgcctggt ttctggccac tgtaggagtg agccccgacc accagggcaa
     4441 gggcttgggc agcgccgtcg tgttgcccgg cgtagaggcc gccgaacgcg ccggtgtgcc
     4501 cgcctttctc gaaacaagcg caccaagaaa ccttccattc tacgagcgcc tgggcttcac
     4561 cgtgaccgcc gatgtcgagg tgcccgaggg acctaggacc tggtgtatga cacgaaaacc
     4621 tggcgcctaa gcggccgcga tccagacatg ataagataca ttgatgagtt tggacaaacc
     4681 acaactagaa tgcagtgaaa aaaatgcttt atttgtgaaa tttgtgatgc tattgcttta
     4741 tttgtaacca ttataagctg caataaacaa gttaacaaca acaattgcat tcattttatg
     4801 tttcaggttc agggggaggt gtgggaggtt ttttaaagca agtaaaacct ctacaaatgg
     4861 ctgatcccaa tggcgcgccg agcttggcgt aatcatggtc atagctgttt cctgtgtgaa
     4921 attgttatcc gctcacaatt ccacacaaca tacgagccgg aagcataaag tgtaaagcct
     4981 ggggtgccta atgagtgagc taactcacat taattgcgtt gcgctcactg cccgctttcc
     5041 agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg
     5101 gtttgcgtat tgggcgctgt tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc
     5161 ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag
     5221 gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa
     5281 aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc
     5341 gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc
     5401 ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg
     5461 cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt
     5521 cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc
     5581 gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc
     5641 cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag
     5701 agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg
     5761 ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa
     5821 ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag
     5881 gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact
     5941 cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa
     6001 attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt
     6061 accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag
     6121 ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca
     6181 gtgctgcaat gataccgcga gaaccacgct caccggctcc agatttatca gcaataaacc
     6241 agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt
     6301 ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg
     6361 ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca
     6421 gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg
     6481 ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca
     6541 tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg
     6601 tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct
     6661 cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca
     6721 tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca
     6781 gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg
     6841 tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac
     6901 ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt
     6961 attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc
     7021 cgcgcacatt tccccgaaaa gtgccacctg acgtctaaga aaccattatt atcatgacat
     7081 taacctataa aaataggcgt atcacgaggc ccttttgtc
//
