We narrowed to 526 results for: sequence

...high-quality portion of a sequencing reaction: Sequence near the beginning or end of a sequencing reaction is often...primer sequence used for each reaction Tips and FAQs My sequence doesn’t match Addgene’s sequencing result... Protocols Sequence Analysis Sequence Analysis of a Plasmid... You can find Addgene's sequencing results by clicking on the "View Sequences" link on the Plasmid Information...but we do not sequence the entire plasmid. Addgene strongly recommends that you sequence any portion ...plasmid backbone (See Addgene's Sequencing Primers for reference). Many sequencing cores have a list of common...obtain each result above the posted sequence in the "View Sequence" link. Is your custom primer found ...

...leader sequence fused to your gene of interest, in frame with the start codon or tag sequences (where ...bp of homologous sequence is used, plus a minimum of 18 bp of your template sequence. 5' and 3' primers...length (typically 10-12 bp) based on a specific sequence. The annealed but nicked vector product is then...at a specified distance from their recognition sequence. This can create multiple distinct overhangs with...final product (no "cloning scars"). A “stuffer” sequence allows for electrophoretic separation of linearized...LIC-specific vector will provide the homologous sequence which must be built into the 5’ end of the respective...primers will have different leader sequences, but operate on the same principle (homologous to the first G...

...8bp). Hybridization Sequence: The region of the primer that binds to the sequence to be amplified (usually...primers for molecular cloning consist of: Leader Sequence: Extra base pairs on the 5' end of the primer ...should use a DNA analysis tool, such as Addgene’s Sequence Analyzer , to allow you to identify which restriction...restriction sites are present in a given sequence. You want to choose enzymes that: Do not cut within your...reverse primer. Next, we need to examine the DNA sequence that we want to amplify and design primers that...reaction. Therefore, our Forward Primer will use the sequence 5'-ATGTGGCATATCTCGAAGTAC-3' for the region that...add TAAGCA, resulting in a final Forward Primer sequence of 5'-TAAGCAGAATTCATGTGGCATATCTCGAAGTAC-3'. For...

...identical sequences on the ends (sequences A and B in the figures). These identical sequences can be created...sites. Join almost any two fragments regardless of sequence. No scar between joined fragments. Fewer steps...segment and a 3′ end that anneals to the target sequence. One strategy is to order primers that are 60 ...adjacent fragment and 30 bp annealing to the target sequence. Avoid strong secondary structures in the homology...correct plasmid product by restriction digest . Sequence the important regions of your final plasmid, particularly...together using oligos When you need intervening sequence between two PCR products, one method is to “stitch...introducing promoters, terminators, and other short sequences into the assembly and is used when the part to...

...DNA sequence (and where they will cut), use a sequence analysis program such as Addgene's Sequence Analyzer...occurring enzymes that cleave DNA at specific sequences. There are hundreds of different restriction enzymes...scientists to target a wide variety of recognition sequences. For a list of many commonly used restriction ...sensitive restriction sites . Sometimes enzymes cut sequences which are similar, but not identical, to their...

...that you select multiple target sequences for each gene. Some sequences will be more effective than others...oligos from section B contain the shRNA sequence flanked by sequences that are compatible with the sticky ...fragment and a 5kb fragment. 5. Sequence positive clones with pLKO.1 sequencing primer (5’ CAA GGC TGT TAG ...I. Appendix I.1. Sequence of pLKO.1 TRC-Cloning Vector Click here to see the sequence of pLKO.1 TRC-cloning...Published articles H.2 Web resources I. Appendix I.1 Sequence of pLKO.1 TRC-Cloning Vector I.2 Recipes I.3 Warranty...destroyed in most cases (depending on the target sequence), while the EcoRI site is preserved. For a complete...downstream of the start codon (ATG), search for 21nt sequences that match the pattern AA(N 19 ). If no suitable...

...endonucleases that recognize a large range of DNA sequences. Given the variety of these enzymes and the unique...that restriction enzymes cleave DNA at specific sequences called restrictions sites. Often, the size of .... Many DNA analysis tools, including Addgene’s Sequence Analyzer , allow you to identify which restriction...restriction sites are present in a given sequence. For a list of the commonly used commercially available restriction...expensive forms of plasmid verification, such as DNA sequencing . In the example above, digestion with enzyme...

... Watch the Video! Sequence Analysis Verify important plasmid features using sequence analysis RNA Extraction...with Lentivirus Genomically integrate your DNA sequence into a cell line Isolating a Monoclonal Cell Population...

...property of primers is that they must correspond to sequences on the template molecule (must be complementary...efficiently (e.g. GCGGCG-restriction site-your sequence). Protocol Video Watch the protocol video below...

...Modify your plasmid MCS or add short sequences using oligo overlap cloning....colonies for mini-prepping and verify insert by sequencing. Reference Page Top Index...

...also stable enough to now bind to the primer DNA sequence. Extend DNA for 1 minute at 72°C: The Taq polymerase...of dH 2 O. Tips and FAQ Tips If your target DNA sequence is GC-rich, increase the time of the denaturing...adjusted according to the length of the target sequence. FAQ How do I design primers? See our reference...

...meningitidis Cas9, or Cas12a from your input sequence or list of sequences. Developed by the George Church Lab ...new window) Identifies gRNA target sequences from an input sequence and checks for off-target binding....window) Identifies candidate gRNA target sequences in an input sequence, which can be an accession number, ..., genomic location, pasted nucleotide sequence, or a sequence text file you upload. Currently supports...program that identifies gRNA target sequences from an input sequence, checks for off-target binding and...window) Identification of gRNA target sequences from an input sequence and checks for off-target binding....

...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...

...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...

... be any ~20 nucleotide sequence, provided it meets two conditions: The sequence is unique compared to ...(PAM) The PAM sequence (NGG) serves as a binding signal for Cas9, but the exact sequence depends on which... target, the seed sequence (8–10 bases at the 3′ end of the gRNA targeting sequence) will begin to anneal...matters. Mismatches between the target sequence in the 3′ seed sequence inhibit target cleavage, while mismatches...determined by both the gRNA sequence and the Cas9 enzyme. Ideally, a gRNA targeting sequence will have perfect ...require PAM sequences in very close proximity to the region to be edited, making the PAM sequence (NGG) limiting...target sequence in your donor DNA with a silent mutation that does not change the amino acid sequence. HDR...

...) Many DNA analysis tools, including Addgene’s Sequence Analyzer , allow you to identify which restriction...restriction sites are present in a given sequence. When selecting restriction enzymes, you want to choose enzymes...

...additional 5' clamping sequence such as GTTTCTTC). Note that all prefix/suffix sequences are given for the ...Assembly standards define standard prefix and suffix sequences that make it easy to combine components for synthetic...sites which preserve a standard prefix and suffix sequence at either end of the assembly, even as more parts...registries, including the specific prefix and suffix sequences for each (these can be built into PCR primers ...prohibited sites and must be omitted from the coding sequence of the part and backbone. For some standards, ...Thr-Gly; Use N-parts prefix to preserve N-term sequence. Prohibited Restriction Sites: EcoRI, XbaI, NgoMIV...

...-MeCP2 fusion for gene and non-coding enhancer sequence repression dCas9-VPR fusion for gene activation...express dCas9-VPR for activation. Usage Guide sequences can be designed using Benchling or using similar...Golden Gate cloning to insert the desired guide RNA sequence by digestion of the backbone with BbsI followed...phosphorylated oligonucleotides containing the guide target sequence: Up to two guides can be expressed from the same... Dual Guide Protocol (PDF, 990 KB) Plasmid and Sequence Derivative Sources Original pX458 backbone ( Addgene...

...Palmitoylation sequence from GAP43 EGFP Connie Cepko 61099 Lck-GFP Membranes Palmitoylation sequence from Lck...4xmts-mNeonGreen Mitochondria 4x Mitochondrial targeting sequence (COX8) mNeonGreen Dorus Gadella 98819 4xmt-mTurquoise2...4xmt-mTurquoise2 Mitochondria 4x Mitochondrial targeting sequence (COX8) mTurquoise2 Dorus Gadella 98818 4xmts-mScarlet-I...4xmts-mScarlet-I Mitochondria 4x Mitochondrial targeting sequence (COX8) mScarlet-I Dorus Gadella 19031 pABCb10-...pLV-mitoGFP Mitochondria Mitochondrial targeting sequence (COX8) GFP Pantelis Tsoulfas 80000 pMito-miRFP703...pMito-miRFP703 Mitochondria Mitochondrial targeting sequence (COX8) miRFP703 Vladislav Verkhusha 36208 pmTurquoise2...pmTurquoise2-Mito Mitochondria Mitochondrial targeting sequence (COX8) mTurquoise2 Dorus Gadella 26673 pEGFP-C1...

... DNA sequence complementary to the crRNA. In Type II CRISPR systems, a potential target sequence is only...Regularly Interspaced Short Palindromic Repeat) sequences were initially discovered in the E. coli genome...loci to generate CRISPR RNAs (crRNAs). Through sequence homology, these crRNAs guide a Cas nuclease to...material, which must also contain a species-specific sequence known as a protospacer adjacent motif (PAM). The...CRISPR array contains many unique protospacer sequences that have homology to foreign DNA. Protospacers...Protospacers are separated by short palindromic repeat sequences. (1) The CRISPR array is transcribed to make the...genomes, are highly diverse, with varying PAM sequences and number/types of Cas proteins. Makarova et ...