Skip to main content
Addgene
Open Menu

Enter a sequence to perform a BLAST-based search of Addgene plasmid sequence databases.

In Category

We narrowed to 538 results for: sequence

Showing: 141 - 160 of 538 results

  1. Plasmids 101: Gateway Cloning

    Type
    Blog Post
    ... a rapid and highly efficient way to move DNA sequences into multiple vector systems. With the appropriate...scientists can easily clone in their desired DNA sequences. Gateway technology relies on the two reactions...add attB sites to either end of the KRAS coding sequence. If you choose this strategy, it’s important to...protein expression elements (ribosome recognition sequences, start codon, stop codons, reading frame considerations...verify the integrity of your expression clone via sequencing or restriction digest! Then, you can transform...Once you generate the entry clone with your DNA sequence of interest, you can move this DNA fragment across...

  2. INTEGRATE: Bacterial Genome Engineering Using CRISPR-Transposons

    Type
    Blog Post
    ...mariner), as a method of tagmentation for next-gen sequencing pipelines (e.g. using Tn5), and as knock-in tools...A CRISPR RNA (crRNA) comprising a 32 nt spacer sequence specifying the target site. Four proteins (TniQ...interest flanked by ~50-150 bp transposon end sequences (R and L ends).  Figure 1: (A) Components.... Genomic targets require an upstream 2-bp PAM sequence for recognition by QCascade; although 5’-CC-3’...complexes survey the genome and bind target DNA sequences using RNA-DNA base pairing. Separately, TnsABC...overnight Screen colonies phenotypically or by PCR/sequencing While CRISPR-transposons are powerful and simple...

  3. CRISPR 101: RNA Editing with Cas13

    Type
    Blog Post
    ... cells. Cas13 enzymes also don’t require a PAM sequence at the target locus, making them more flexible...given single base protospacer flanking site (PFS) sequence, but orthologs like LwaCas13a do not require a... target RNA from bases 12-26 of a 30 nt target sequence. For the editing portion of the protein, they ...designated this system REPAIRv1. Using next-generation sequencing, they confirmed A->I editing and found that 50...target effects observed in whole-transcriptome RNA-sequencing, the authors performed rational mutagenesis and...RESCUE further expands the RNA editing toolkit and sequences that can be edited.   As discussed above, the ...

  4. Avoiding the Dark Side of Fluorescent Protein Fusions with mOX FPs

    Type
    Blog Post
    ...positioning of the FP sequence relative to cell compartment targeting sequences, which often have absolute...requirements. For example, a KDEL ER retrieval sequence only functions at the extreme C-terminus [3]. ...Additionally, FPs that encode N-glycosylation consensus sequences are modified with the addition of N-glycans [9...using FPs containing two cysteine residues and a sequence intended to localize them to the secretory pathway...cysteine residues and N-glycosylation consensus sequences in popular FPs. This generated a palette of inert...others (5, 6, 7, 10, 11) highlight the very real consequences of misfolded dark FP fusions including erroneous...

  5. Five Popular Model Organisms, Part 2

    Type
    Blog Post
    ...this gap, the Rat Genome Sequencing Project conducted a large scale sequencing and concluded that nearly... fully sequenced by the year 2000, becoming the first plant to have its entire genome sequenced (Koornneff... limited at first, advances in axolotl genome sequencing have vastly expanded axolotl research. One of...of the most significant advances is the sequencing of the entire axolotl genome, revealing an abundance..., R., Weinstock, G., Metzker, M. et al. Genome sequence of the Brown Norway rat yields insights into mammalian...

  6. Plan Your Experiment

    Type
    Collection
    ...perfect world, your gRNA sequence would be an exact complement to your target sequence with no homologous sites... the desired target sequence, or on-target activity. Two gRNA targeting sequences with 100% homology to... selected target sequence. Additionally, any mismatches between a given gRNA sequence and your genomic...customization Usually paired with a sequencing method Can’t provide specific sequence information alone Digital ...rare edits Many options like RNA-sequencing, Perturb-seq, deep sequencing, etc. qPCR Knockouts Activation...and the guide RNA (gRNA), the design of the gRNA sequence, and finally how to validate your genetic edit...within a very narrow window proximal to the PAM sequence Base Edit Prime Edit Generate targeted insertions...

  7. A Practical Guide to Optimizing AAV DIO and FLEx Vector Expression

    Type
    Blog Post
    ...the same sequence to flank a transgene (homologous pairs) or used different scrambled sequences to flank...Recombinase-dependent vectors utilize pairs of specific sequences, lox (recognized by Cre) or frt (recognized by...used a lox-STOP-lox (LSL) or frt-STOP-frt (FSF) sequence prior to an ATG start codon to prevent transcription...recognized by Cre, but they are comprised of different sequences (loxp and lox2272) and can not recombine with ...phenomena by analyzing the plasmid and viral DNA sequences and expression profile at multiple steps from ...containing a DIO/FLEx cassette using next-generation sequencing (NGS) (Guerin, K. 2020). In general, we observe...Mitsopoulos A, Fan M (2020) A Novel Next-Generation Sequencing and Analysis Platform to Assess the Identity ...

  8. 28 Hot Plasmid Technologies from 2015

    Type
    Blog Post
    ...initiation sequence known as the Species-Independent Translational Sequence (SITS). This sequence bypasses... via DNA sequencing or other readout. Cellular memory is accomplished by the use of sequence-specific ...in series with unique spacer sequences (pMemoryArray) gives a DNA sequence capable of recording 11 bits...shuttled out of the nucleus due to the nuclear export sequence. Treatment with rapamycin induces Cas9(N)-FRB-... the presence of multiple copies of targeting sequences. Scientists have adopted and employed this concept...inducible expression, loxP sites, H2B, P2A or PEST2D sequences. Several pre-constructed promoter-selection marker...generating plasmids with varying numbers of repeated sequences or inserts, which can be advantageous for imaging...

  9. Pairing CombiGEM and CRISPR for Combinatorial Genetic Screening

    Type
    Blog Post
    ...gRNA targeting sequences, you can then readily generate your barcoded gRNA library sequences via oligo synthesis...can be tracked and quantitatively analyzed by sequencing its set of barcodes. CombiGEM is highly flexible...will need is to get the list of effective gRNA sequences targeting your genes of interest. Thanks to the...where NNN and XXX represent the 20-bp gRNA target sequence and its 8-bp barcode, respectively. Once cloned...

  10. Multiplex Genome Editing with CRISPR-Cpf1

    Type
    Blog Post
    ...regulatory sequences to allow for expression of multiple gRNAs (i.e. Csy4 cleavage sequence, tRNAs, multiple...No dsDNA cleavage Blunt end 5' overhang PAM sequence NGG TTTV PAM site preserved? Usually Destroyed...multiple promoters to drive crRNA expression and sequences that allow for array processing, i.e. tRNAs or...Lentiviruses carry a (+) strand RNA copy of the DNA sequence, which is a suitable substrate for Cpf1. This ...

  11. Plasmids for Endogenous Gene Tagging in Human Cells

    Type
    Blog Post
    ...side of the fluorescent protein sequence that is homologous to the sequence into which it is being inserted...Gene editing is the addition of a useful coding sequence (i.e. GFP) into a gene while leaving other features...in one edited allele, or gene copy. The coding sequence for GFP is transcribed and translated along with...the gene of interest, a linker, and the GFP tag sequence to be inserted. Created with BioRender.com. ...

  12. What's New in CRISPR - November 2021

    Type
    Blog Post
    ...adapted the native PEG10 RNA sequences to instead drive packaging of cargo sequences in the capsid, in an approach...also optimized the architecture and amino acid sequence of the prime editor, yielding PEmax, to work synergistically...plasmids  Deletion and replacement of long genomic sequences The PEDAR strategy engineered by Wen Xue’s lab...double-strand breaks and deletes the intervening sequence. As with other prime editing tools, the desired...

  13. 10 Great Guest Posts We're Thankful For

    Type
    Blog Post
    ...described in this post are engineered to bind RNA sequences derived from Zika virus and, upon binding, produce...stripped down viral genomes containing only the sequences required for viral replication and transcription...that can be used to quickly barcode a set of DNA sequences. In this post, Alan Wong (formerly of the Lu Lab... variety of customizable functions to your DNA sequence of interest. Potential applications of the system...

  14. Plasmids 101: What is a plasmid?

    Type
    Blog Post
    ...single-stranded DNA sequence used as an initiation point for PCR amplification or sequencing. Primers can be... Description Origin of Replication (ORI) DNA sequence which allows initiation of replication within ... be exploited for sequence verification of plasmids.   How is a plasmid constructed in the lab? Due...

  15. When Fidelity Matters: A frank discussion about ligase fidelity

    Type
    Blog Post
    ...cloning), adaptor ligation for next generation sequencing (NGS) library construction, and circularization...These applications detect specific nucleotide sequences with high specificity and quantitative accuracy...Detection Reaction (LDR), probes complementary to the sequence of interest are annealed to target DNA (genomic...fidelity thermostable DNA ligase. If the target sequence is present, the probes will ligate. Subsequent... contacts, and thus do not read specific base sequences, but are sensitive to distortions in helix shape...Gelfand. "Cloning, overexpression and nucleotide sequence of a thermostable DNA ligase-encoding gene." Gene...1756968. 7. Lauer, G., et al. "Cloning, nucleotide sequence, and engineered expression of Thermus thermophilus...

  16. An “elegans” Approach to Better CRISPR/Cas9 Editing Efficiency

    Type
    Blog Post
    ...injected animal.  PAM sequence considerations For changes 5’ to the PAM, sequences corresponding to the...developmental processes, CRISPR/Cas9 allows us to delete sequences and introduce mutations and epitopes with unprecedented... but did result in 1-2 kilobases of additional sequence being introduced. Cre-mediated excision of the...the selective cassette minimizes the sequence added, leaving a 34 bp “scar”, but currently requires additional...editing efficiency, while for edits 3’ to the PAM sequences corresponding to the opposite strand were better...they were able to insert 10 kb of non-homologous sequence or a series of single-nucleotide substitutions...an alternative gene editing system for AT-rich sequences https://doi.org/10.17912/W2237D Farboud B, Severson...

  17. Technique: Probe Phage Genomes for Host Binding Proteins

    Type
    Blog Post
    ...the need for genome sequencing or annotation (5). In the event that genome sequencing can be done, this ...problem: Host binding proteins are hard to find Sequencing and cloning phage DNA can be difficult, in part...(4). To further complicate matters, even when sequenced, phage genomes tend to be choc-full of open reading...technique can also help pinpoint the genes within a sequenced genome that encode for functional cell-binding... a restriction enzyme with a short recognition sequence (like MluCI) that will cleave DNA frequently enough...Now you can grow up your colonies of interest, sequence the inserted DNA and characterize your new putative...

  18. CRISPR 101: Homology Directed Repair

    Type
    Blog Post
    ...insertable sequences or specific mutations, along with homologous DNA to the target sequence to be modified...and deletions along the way. HDR uses homologous sequence as a template for repair and is thus the more ...used for repair is identical to the original DNA sequence at the DSB, or it can introduce specific mutations...non-crossover events. This means all newly synthesized sequences are retained by the original damaged DNA molecule...is introduced disrupting one or both of these sequences. To prevent re-cutting and the resulting mutations...

  19. Quickest Way to Deposit Plasmids: The Deposit Spreadsheet

    Type
    Blog Post
    ... also directly paste sequence data into the appropriate full and partial sequence cells. *Note: If you...strain DH5α. For plasmids with highly repetitive sequences (prone to recombination in bacteria) such as lentiviral...username. Shipping address and phone number. Plasmid sequences or GenBank files. We encourage submission of QUEEN-generated...GenBank files (Mori and Yachie, 2021) but can accept sequence files in any format. Distribution status for your... amino acid change as well as any functional consequences of that change. For example, “Mutated aspartate...

  20. CRISPR Plasmids - RNA Targeting

    Type
    Collection
    ...target have high sequence identity to the spacer sequence and contain a PAM sequence just downstream of...CRISPR plasmids for targeting RNA sequences, which include the enzymes C2c2/Cas13a and Cas13b. CRISPR...they have recognized and cleaved the target RNA sequence, they adopt an enzymatically active state and ... of the sequence to be cleaved. This non-specific cleavage is thought to activate programmed cell death...
Showing: 141 - 160 of 538 results