![]() "Recombinase"-independent recombineering.FAQ 14: Why don’t I get the high frequencies that you report for ssDNA oligo recombineering?.FAQ 13: What do I do with the tube containing the “gel-like stuff” in it when you send me a strain or plasmid?.FAQ 12: How should I maintain the recombineering strains?.FAQ 11: Is the amount of NaCl in the LB medium important?.FAQ 10: Why do my electroporations “pop” (arc)?.FAQ 9: What conditions should I use for electroporation?.FAQ 8: I get tons of recombinants but when I analyze them, none are in the right place.FAQ 7: Why don’t I get any recombinants?.FAQ 6: What happens if I induce for longer or shorter times?.FAQ 5: Do the oligos I order need to be purified?.FAQ 4: Can’t I use an air shaker for the 42° induction?.FAQ 3: What happens if I grow the recombineering cells at temperatures higher than 32☌?.FAQ 2: What sequences do I use to make a drugR cassette for knock-outs?. ![]() FAQ 1: How much antibiotic should I use for chromosomal/BAC (single-copy) or plasmid (multi-copy) constructs?.“Recombinase”-independent recombineering.On the mechanism of ssDNA recombination.This will determine whether or not it will be read correctly. In addition, the combination of restriction enzymes you use determines how a genetic sequence will fit in a construct. This is of particular importance because you don’t want to use an enzyme that can cut somewhere in the middle of a genetic sequence. There are a number of things to consider when applying restriction cloning, such as which restriction enzymes to use. SnapGene allows you to simulate this process either by using the protocols it has available or creating your own. This process is performed in the lab following a given protocol. A number of cut sequences can be joined together with a process known as ligation. Restriction cloning is a method of editing genetic sequences by cutting them with restriction enzyme at suitable restriction sites. The Sequence view also shows the features seen in the map view, and adds an extra feature, a red mark that indicates the stop codons of your sequence. The Sequence view shows the characters in the sequence is convenient when editing sequences because it allows you to pinpoint the critical locations in the sequence. This information is important for estimating the cost of synthesizing your sequence and is useful in a number of calculations. This includes the number of base pairs, the number of amino acids and the molecular weight of the sequence in Daltons. The Features view, which is accessed by clicking on Views and selecting Features, shows some information about the sequences you have. These views allow you to see features included in your file, the restriction enzymes in your sequence, the actual characters in the sequence, and your primers. Other views can be accessed by selecting one of the options in the Views menu. If the length and direction of the reading frames feature are the same as the length and direction of your gene’s feature, the gene will likely be transcribed correctly. These features allow you to see the reading frames of the sequences you have in your file, and highlights the parts of your construct that hold given sequences.Ĭlicking on show translate on the left adds some features that show the reading frames of each sequence. ![]() The default view is the map which hides the letters in the genetic sequence to create space for visual features. There are a number of ways you can view a genetic sequence in SnapGene. This is extremely useful in a field like synthetic biology where existing sequences are combined to form new ones that allow organisms to perform novel functions. SnapGene is software that allows you to visualize and edit genetic sequences. ![]()
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