Basic Vector Information
Expresses the 5-HT sensor GRAB_5-HT1.0 in neurons, could be used as a genetically encoded sensor for measuring serotonin dynamics
pAAV-hsyn-GRAB_5-HT1.0 vector Vector Map
Plasmid Resuspension Protocol:
1. Centrifuge at 5,000×g for 5 min.
2. Carefully open the tube and add 20 μl of sterile water to dissolve the DNA.
3. Close the tube and incubate for 10 minutes at room temperature.
4. Briefly vortex the tube and then do a quick spin to concentrate the liquid at the bottom. Speed is less than 5000×g.
5.Store the plasmid at -20 ℃.
pAAV-hsyn-GRAB_5-HT1.0 vector Sequence
LOCUS pAAV-hsyn-GRAB_5 6530 bp 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) TITLE Direct Submission REFERENCE 3 (bases 1 to 6530) AUTHORS . TITLE Direct Submission COMMENT SGRef: number: 1; type: "Journal Article" COMMENT SGRef: number: 2; type: "Journal Article" FEATURES Location/Qualifiers source 1..6530 /mol_type="other DNA" /organism="synthetic DNA construct" repeat_region 1..130 /label=AAV2 ITR (alternate) /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 /label=Kozak sequence /note="Kozak sequence" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" /regulatory_class="other" CDS 587..649 /codon_start=1 /product="leader sequence from mouse immunoglobulin kappa light chain" /label=leader sequence from mouse immunoglobulin kappa /note="Ig-kappa leader" /translation="METDTLLLWVLLLWVPGSTGD" protein_bind 650..674 /gene="mutant version of attB" /label=BP Clonase(TM) binding site /bound_moiety="BP Clonase(TM)" /note="attB1" /note="recombination site for the Gateway(R) BP reaction" CDS 683..1420 /codon_start=1 /label=Human HTR2C(2-247) /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 1451..1702 /label=VC155 /note="C-terminal fragment of mVenus for use in bimolecular fluorescence complementation (BiFC) (Kodama and Hu, 2010)" CDS 2177..2644 /codon_start=1 /label=Human HTR2C(304-458) /note="Human HTR2C(304-458)" /translation="NNERKASKVLGIVFFVFLIMWCPFFITNILSVLCEKSCNQKLMEK LLNVFVWIGYVCSGINPLVYTLFNKIYRRAFSNYLRCNYKVEKKPPVRQIPRVAATALS GRELNVNIYRHTNEPVIEKASDNEPGIEMQVENLELPVNPSSVVSERISSV" misc_feature 2657..3245 /label=WPRE /note="woodchuck hepatitis virus posttranscriptional regulatory element" polyA_signal 3277..3753 /label=hGH poly(A) signal /note="human growth hormone polyadenylation signal" repeat_region 3793..3933 /label=AAV2 ITR /note="inverted terminal repeat of adeno-associated virus serotype 2" rep_origin 4008..4463 /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" promoter 4745..4849 /label=AmpR promoter CDS 4850..5707 /label=AmpR /note="beta-lactamase" rep_origin 5881..6469 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication"
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