LOCUS       sequence-207616-        5406 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Homology arms and mEGFP-linker sequence for N-terminus tagging of 
            human RAB5A.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5406)
  AUTHORS   Allen Institute for Cell Science
  TITLE     Allen Institute for Cell Science Plasmids
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 5406)
  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: 
            "Unpublished"
COMMENT     Sequence Label: AICSDP-39:RAB5A-mEGFP
FEATURES             Location/Qualifiers
     source          1..5406
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     173..195
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     187..204
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     188..204
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             join(1222..1224,1225..1227,1228..1938)
                     /codon_start=1
                     /product="enhanced GFP with monomerizing A206K mutation 
                     (Zacharias et al., 2002)"
                     /label=mEGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1267..1288)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1528..1547)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1875..1896
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(2994..3010)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2994..3010)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3007..3029)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3018..3034
                     /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(3042..3048,3049..3066,3067..3072))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3087..3108
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3225..3242)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3396..3984)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3476..3495)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4155..4946,4947..5015))
                     /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     4778..4797
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5016..5120)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5188..5206
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(5244..5266)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     5366..5385
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 tgtgaaatac cgcacagatg cgtaaggaga aaataccgca tcaggcgcca ttcgccattc
       61 aggctgcgca actgttggga agggcgatcg gtgcgggcct cttcgctatt acgccagctg
      121 gcgaaagggg gatgtgctgc aaggcgatta agttgggtaa cgccagggtt ttcccagtca
      181 cgacgttgta aaacgacggc cagtgaattg acgcgtattg ggatgttctg agcgcctgta
      241 gtctcagcta ctcaggaggt tgaggcagga ggattgcttg agcccaggag gtctaggttg
      301 cagtgagctg tgaccacgcc actgcactcc agcctgggtg agagagcaaa accctttctc
      361 caaaaaaatt tttttaaaaa tggcatgtat attgttcgtc taaagactac aaaaaagtta
      421 caattattgt aagtaaataa gtcagtaata atcaccacat ggagaacaac tgttagcatt
      481 tctgtgtatg gtgtgcttga gcagatgagg aaacagtttt aagtctgccg ttaggcacac
      541 aaacctggga gactgaaaat atttgttgtc caaaattacc attaattaaa tgaaaaatgc
      601 atcgtttaac agaaaacttc aaataatact aaattttagg atctgttcta aaagtttccc
      661 atttattaat aacataattt ttctcaaaac aatgttaatc caagtagcct ggctacagaa
      721 cttgaactcg agtcatctct caggttgcct gggtttgaaa ctcggctctg tcacttctgt
      781 gacaataact gtgtgacctc aattaagctt cttaacatct ctgtgcttgt ttcctcattt
      841 ataaggattg taataatatc tcattttata ggattgttga gaagttaagt acttaaagca
      901 ctgcctggca caaacttgaa gctttgaggt tagcctttaa aaaaaaaagt aattttggac
      961 tcacatccta tacctttgaa tttcctttaa aagatagtga cataatctaa agggcattga
     1021 aggtctagta gagtttaaga taatcatgct ataggcaata tttaaagtga ttaatagtaa
     1081 atttgcttta ctgatttgta tatttaattc ataattgttt ctttacaggt ttctttacct
     1141 ccagaaagaa gaatattggc cccttgaatt ctggaagttc attgaagagt ctgaaattag
     1201 ggacttattt caaatttgga catggtgagc aagggcgagg agctgttcac cggggtggtg
     1261 cccatcctgg tcgagctgga cggcgacgta aacggccaca agttcagcgt gtccggcgag
     1321 ggcgagggcg atgccaccta cggcaagctg accctgaagt tcatctgcac caccggcaag
     1381 ctgcccgtgc cctggcccac cctcgtgacc accctgacct acggcgtgca gtgcttcagc
     1441 cgctaccccg accacatgaa gcagcacgac ttcttcaagt ccgccatgcc cgaaggctac
     1501 gtccaggagc gcaccatctt cttcaaggac gacggcaact acaagacccg cgccgaggtg
     1561 aagttcgagg gcgacaccct ggtgaaccgc atcgagctga agggcatcga cttcaaggag
     1621 gacggcaaca tcctggggca caagctggag tacaactaca acagccacaa cgtctatatc
     1681 atggccgaca agcagaagaa cggcatcaag gtgaacttca agatccgcca caacatcgag
     1741 gacggcagcg tgcagctcgc cgaccactac cagcagaaca cccccatcgg cgacggcccc
     1801 gtgctgctgc ccgacaacca ctacctgagc acccagtcca agctgagcaa agaccccaac
     1861 gagaagcgcg atcacatggt cctgctggag ttcgtgaccg ccgccgggat cactctcggc
     1921 atggacgagc tgtacaagag cggcctgaga agcagggccg ctagtcgagg cgccacacgg
     1981 ccaaacgggc caaatacggg aaataaaata tgccagttca aactagtact tctgggagag
     2041 tccgctgttg gcaaatcaag cctagtgctt cgttttgtga aaggccaatt tcatgaattt
     2101 caagagagta ccattggggg tgagattttc ttttttccct gcttcattta atcgaatcat
     2161 acattgacag aaaatatttt gatgttccaa ttgtgtgatt atgaatagct aaataagtca
     2221 tagagtgaag aaaagagctg ttcagcttac cttagcttta aaacttgcct ttatctttgt
     2281 taaaatgatg tgtttttata tttgattgtt tggttaatcc cttgtggtta tattgtagct
     2341 aaaatacaga taaaaaacat gagtagaatc agtgtctcaa gtaataacat ttgacctgat
     2401 acctatttct agctaacaac tagaagcttc ttcaaattaa tacagtacat gtttgtgctc
     2461 aattatattg gtttaaaatg aatagtaaaa aaggtaacct taaattctga tctatataac
     2521 tgaagtattt gcaaataatt atacttactt gtaaaattaa atgtaattaa atactgctag
     2581 atgtactttt ttttggtatt taagagatgg gatcttgctg tcgcctaggc tggagtgcag
     2641 tgtccggatc atagctcact gcagactgca gcctgaaact ctaggctcaa atgatcttct
     2701 tgccttagcc tcccaccaag tagttggacc taagggtgca tgccaccatg ccaggcaagt
     2761 tttttttttt tttttttaag agtcagagtc tcgctgtgtt tccaggttgg tcttgaactc
     2821 ctggcctcaa gtgagcctcc agcctcaacc tcccaggtag ctgtgattac aggtgctccc
     2881 tgctgagatg catatgtaag aatgctccta agctgaggga agagaaaaag tgattaatgg
     2941 aaagggacat aacaggaaaa tcccaatggc gcgccgagct tggcgtaatc atggtcatag
     3001 ctgtttcctg tgtgaaattg ttatccgctc acaattccac acaacatacg agccggaagc
     3061 ataaagtgta aagcctgggg tgcctaatga gtgagctaac tcacattaat tgcgttgcgc
     3121 tcactgcccg ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa
     3181 cgcgcgggga gaggcggttt gcgtattggg cgctcttccg cttcctcgct cactgactcg
     3241 ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg
     3301 ttatccacag aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag
     3361 gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac
     3421 gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga
     3481 taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt
     3541 accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc
     3601 tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc
     3661 cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta
     3721 agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat
     3781 gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca
     3841 gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct
     3901 tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt
     3961 acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct
     4021 cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc
     4081 acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa
     4141 acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta
     4201 tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc
     4261 ttaccatctg gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat
     4321 ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta
     4381 tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt
     4441 aatagtttgc gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt
     4501 ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg
     4561 ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc
     4621 gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc
     4681 gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg
     4741 cggcgaccga gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga
     4801 actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta
     4861 ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct
     4921 tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag
     4981 ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca atattattga
     5041 agcatttatc agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat
     5101 aaacaaatag gggttccgcg cacatttccc cgaaaagtgc cacctgacgt ctaagaaacc
     5161 attattatca tgacattaac ctataaaaat aggcgtatca cgaggccctt tcgtctcgcg
     5221 cgtttcggtg atgacggtga aaacctctga cacatgcagc tcccggagac ggtcacagct
     5281 tgtctgtaag cggatgccgg gagcagacaa gcccgtcagg gcgcgtcagc gggtgttggc
     5341 gggtgtcggg gctggcttaa ctatgcggca tcagagcaga ttgtactgag agtgcaccat
     5401 atgcgg
//
