LOCUS       Exported                6530 bp ds-DNA     circular SYN 09-APR-2021
DEFINITION  Expresses the 5-HT sensor GRAB_5-HT1.0 in neurons.
ACCESSION   .
VERSION     .
KEYWORDS    pAAV-hsyn-GRAB_5-HT1.0
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6530)
  TITLE     Yulong Li 5-HT sensor and DA sensor plasmids
REFERENCE   2  (bases 1 to 6530)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..6530
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     repeat_region   1..130
                     /note="AAV2 ITR (alternate)"
                     /note="Functional equivalent of wild-type AAV2 ITR"
     promoter        145..586
                     /note="synapsin"
                     /note="human synapsin promoter is a neuron-specific 
                     promoter"
     regulatory      581..590
                     /regulatory_class="other"
                     /note="Kozak sequence"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     CDS             587..649
                     /codon_start=1
                     /product="leader sequence from mouse immunoglobulin kappa 
                     light chain"
                     /note="Ig-kappa leader"
                     /translation="METDTLLLWVLLLWVPGSTGD"
     protein_bind    650..674
                     /gene="mutant version of attB"
                     /bound_moiety="BP Clonase(TM)"
                     /note="attB1"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             683..1420
                     /codon_start=1
                     /note="Human HTR2C(2-247)"
                     /note="G-protein coupled receptor for 5-hydroxytryptamine 
                     (serotonin). Also functions as a receptor for various drugs
                     and psychoactive substances, including ergot alkaloid 
                     derivatives, 1-2,5,-dimethoxy-4-iodophenyl-2-aminopropane 
                     (DOI) and lysergic acid diethylamide (LSD). Ligand binding 
                     causes a conformation change that triggers signaling via 
                     guanine nucleotide-binding proteins (G proteins) and 
                     modulates the activity of down-stream effectors. 
                     Beta-arrestin family members inhibit signaling via G 
                     proteins and mediate activation of alternative signaling 
                     pathways. Signaling activates a 
                     phosphatidylinositol-calcium second messenger system that 
                     modulates the activity of phosphatidylinositol 3-kinase and
                     down-stream signaling cascades and promotes the release of 
                     Ca2+ ions from intracellular stores. Regulates neuronal 
                     activity via the activation of short transient receptor 
                     potential calcium channels in the brain, and thereby 
                     modulates the activation of pro-opiomelacortin neurons and 
                     the release of CRH that then regulates the release of 
                     corticosterone. Plays a role in the regulation of appetite 
                     and eating behavior, responses to anxiogenic stimuli and 
                     stress. Plays a role in insulin sensitivity and glucose 
                     homeostasis."
                     /translation="VNLRNAVHSFLVHLIGLLVWQCDISVSPVAAIVTDIFNTSDGGRF
                     KFPDGVQNWPALSIVIIIIMTIGGNILVIMAVSMEKKLHNATNYFLMSLAIADMLVGLL
                     VMPLSLLAILYDYVWPLPRYLCPVWISLDVLFSTASIMHLCAISLDRYVAIRNPIEHSR
                     FNSRTKAIMKIAIVWAISIGVSVPIPVIGLRDEEKVFVNNTTCVLNDPNFVLIGSFVAF
                     FIPLTIMVITYCLTIYVLRRQALM"
     CDS             1433..2161
                     /codon_start=1
                     /note="circular-permutated EGFP(cpEGFP)"
                     /note="When 5-HT binds to HTR2C, conformational changes due
                     to the HTR2C-5-HT interaction induce a subsequent 
                     conformational change in cpEGFP, so that the fluorescence 
                     intensity changes."
                     /translation="NVYIKADKQKNGIKANFHIRHNIEDGGVQLAYHYQQNTPIGDGPV
                     LLPDNHYLSVQSKLSKDPNEKRDHMVLLEFVTAAGITLGMDELYKGGTGGQESMSSKGE
                     ELFTGVVPILVELDGDVNGHKFSVSGEGEGDATTGKLTLKFICTTGKLPVPWPTLVTTL
                     TYGVQCFSRYPDHMKQHDFFKSAMPEGYIQERTIFFKDDGNYKTRAEVKFEGDTLVNRI
                     ELKGIDFKEDGNILGHKLEYN"
     CDS             2177..2644
                     /codon_start=1
                     /note="Human HTR2C(304-458)"
                     /translation="NNERKASKVLGIVFFVFLIMWCPFFITNILSVLCEKSCNQKLMEK
                     LLNVFVWIGYVCSGINPLVYTLFNKIYRRAFSNYLRCNYKVEKKPPVRQIPRVAATALS
                     GRELNVNIYRHTNEPVIEKASDNEPGIEMQVENLELPVNPSSVVSERISSV"
     misc_feature    2657..3245
                     /note="WPRE"
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     CDS             complement(3128..3139)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /note="Factor Xa site"
                     /translation="IEGR"
     polyA_signal    3277..3753
                     /note="hGH poly(A) signal"
                     /note="human growth hormone polyadenylation signal"
     repeat_region   3793..3933
                     /note="AAV2 ITR"
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   3793..3922
                     /note="AAV2 ITR"
     rep_origin      4008..4463
                     /direction=RIGHT
                     /note="f1 ori"
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     promoter        4745..4849
                     /gene="bla"
                     /note="AmpR promoter"
     CDS             4850..5710
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /note="AmpR"
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     rep_origin      5881..6469
                     /direction=RIGHT
                     /note="ori"
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"