Difference between revisions of "Part:BBa K3697003"

Revision as of 23:43, 23 October 2020

Homology Arms for KanR integration in B. Subtilis

This part is derived from a portion of the pOpen Yeast plasmid between the AarI and MspA1I cut sites. More information about pOpen Yeast and how to get it through Stanford Free Genes can be found on their website and at this link: https://stanford.freegenes.org/products/popen_build. This sequence was then divided into two homology arms (homology arm 1 and homology arm 2 as marked in the annotations for the part) which should flank the region in the genome where the user would like the recombination to occur. When incorporated into the B. subtilis genome, these two sequences (homology arm 1 and homology arm 2) become the two homology arms needed to trigger B. subtilis' natural process of recombination.

One important thing to not about these homology arms is that they are both 550 base pairs in length. This is important because when trying to trigger a recombination event in B. subtilis it is best to use 2 regions of homology of at least 500 base pairs (totaling at least 1000 base pairs of homology) flanking the region where you would like recombination to occur. Shorter regions of homology can be used sometimes, but some have documented reduced transformation efficiency [1].

Having known homology arms in the genome can be helpful for a couple reasons. One, they could be incorporated into the B. subtilis genome so that any target sequence flanked by regions of high homology to these homology arms could be incorporated into the B. subtilis genome. Two, these homology arms could be incorporated into the B. subtilis genome flanking a sequence of interest, then if the target sequence is taken up by B. subtilis (the portion of the pOpen Yeast plasmid between the AarI and MspA1I cut sites) the sequence being flanked by these homology arms could be excised from the B. subtilis genome.

Sequence and Features

Assembly Compatibility:

10
COMPATIBLE WITH RFC[10]
12
COMPATIBLE WITH RFC[12]
21
COMPATIBLE WITH RFC[21]
23
COMPATIBLE WITH RFC[23]
25
COMPATIBLE WITH RFC[25]
1000
COMPATIBLE WITH RFC[1000]

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 <partinfo>BBa_K3697003 short</partinfo>
-This part is derived from a portion of the pOpen Yeast plasmid between the AarI and MspA1I cut sites. More information about pOpen Yeast and how to get it through Stanford Free Genes can be found on their website and at this link: https://stanford.freegenes.org/products/popen_build. This sequence was then divided into two homology arms (homology arm 1 and homology arm 2 as marked in the annotations for the part). When incorporated into the B. subtilis genome, these two sequences (homology arm 1 and homology arm 2) become the two homology arms needed to trigger B. subtilis' natural process of recombination.
+This part is derived from a portion of the pOpen Yeast plasmid between the AarI and MspA1I cut sites. More information about pOpen Yeast and how to get it through Stanford Free Genes can be found on their website and at this link: https://stanford.freegenes.org/products/popen_build. This sequence was then divided into two homology arms (homology arm 1 and homology arm 2 as marked in the annotations for the part) which should flank the region in the genome where the user would like the recombination to occur. When incorporated into the B. subtilis genome, these two sequences (homology arm 1 and homology arm 2) become the two homology arms needed to trigger B. subtilis' natural process of recombination.
+One important thing to not about these homology arms is that they are both 550 base pairs in length. This is important because when trying to trigger a recombination event in B. subtilis it is best to use 2 regions of homology of at least 500 base pairs (totaling at least 1000 base pairs of homology) flanking the region where you would like recombination to occur. Shorter regions of homology can be used sometimes, but some have documented reduced transformation efficiency [1].
+Having known homology arms in the genome can be helpful for a couple reasons. One, they could be incorporated into the B. subtilis genome so that any target sequence flanked by regions of high homology to these homology arms could be incorporated into the B. subtilis genome. Two, these homology arms could be incorporated into the B. subtilis genome flanking a sequence of interest, then if the target sequence is taken up by B. subtilis (the portion of the pOpen Yeast plasmid between the AarI and MspA1I cut sites) the sequence being flanked by these homology arms could be excised from the B. subtilis genome.
 <!-- Add more about the biology of this part here
 ===Usage and Biology===
-One important thing to not about these homology arms is that they are both 550 base pairs in length. This is important because when trying to trigger a recombination event in B. subtilis it is best to use 2 regions of homology of at 500 base pairs (totaling at least 1000 base pairs of homology). Shorter regions of homology can be used sometimes, but some have documented reduced transformation efficiency[1].
-Having known homology arms in the genome can be helpful for a couple reasons. One, they could be incorporated into the B. subtilis genome so that any target sequence flanked by regions of high homology to these homology arms could be incorporated into the B. subtilis genome. Two, these homology arms could be incorporated into the B. subtilis genome flanking a sequence of interest, then if the target sequence is taken up by B. subtilis (the portion of the pOpen Yeast plasmid between the AarI and MspA1I cut sites) the sequence being flanked by these homology arms could be excised from the B. subtilis genome.
 [1] Dubnau D. Sonenshein AL,  Hoch JA,  Losick R. Genetic exchange and homologous recombination, Bacillus subtilis and Other Gram-positive Bacteria: Biochemistry, Physiology, and Molecular Genetics, 1993Washington, DCASM(pg. 555-584)