Team:CCU Taiwan/Parts

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Overview Basic Parts Composite Parts

Overview

There are total 25 submitted parts in our project, which including basic parts and composite parts. Our project is about the application of CRISPR, thus most of our parts are related to the CRISPR-Cas12a system. Some of them are for the purification of LbCas12a protein, like 10X His (BBa_K2927002), MBP (BBa_K2927003), and TEV site (BBa_K2927004). Some of them are DNA template which is for the purpose of crRNA transcription (BBa_K2927010, BBa_K2927011, BBa_K2927012, BBa_K2927013, BBa_K2927014, BBa_K2927015). In addition, to test whether our purified LbCas12a protein can function properly, there are some mutated crRNA (BBa_K2927020, BBa_K2927021, BBa_K2927022, BBa_K2927023, BBa_K2927024, BBa_K2927025) which have been made a change on 5 nucleotides of crRNA from 5’ end for negative control and a basic part for positive control (BBa_K2927031).

Part number	Description	Supporting information	Length (bp)
BBa_K1614000	T7 Promoter	RNA polymerase binding	18
BBa_K2927001	RBS	Ribosome binding site	23
BBa_K2927002	10X His	For Ni2+ column	30
BBa_K2927003	MBP (maltose binding protein)	For MBP trap column	1101
BBa_K2927004	Tobaco Etch Virus (TEV) motif	TEV protein cutting site	21
BBa_K2927005	LbCas12a	Produce LbCas12a protein	3684
BBa_K395601	T7 Terminator	RNA polymerase leaving	48
BBa_K2927006	guide RNA	guide LbCas12a protein for binding target sequence	12
BBa_K2927010	DNA template of crRNA 1 for LbCas12a	DNA of ASFV for the detection	24
BBa_K2927011	DNA template of crRNA 2 for LbCas12a	DNA of ASFV for the detection	24
BBa_K2927012	DNA template of crRNA 3 for LbCas12a	DNA of ASFV for the detection	24
BBa_K2927013	DNA template of crRNA 4 for LbCas12a	DNA of ASFV for the detection	24
BBa_K2927014	DNA template of crRNA 5 for LbCas12a	DNA of ASFV for the detection	24
BBa_K2927015	DNA template of crRNA 6 for LbCas12a	DNA of ASFV for the detection	24
BBa_K2927020	Negative control sequence for crRNA 1 target	negative control for CRISPR-LbCas12a system	28
BBa_K2927021	Negative control sequence for crRNA 2 target	negative control for CRISPR-LbCas12a system	28
BBa_K2927022	Negative control sequence for crRNA 3 target	negative control for CRISPR-LbCas12a system	28
BBa_K2927023	Negative control sequence for crRNA 4 target	negative control for CRISPR-LbCas12a system	28
BBa_K2927024	Negative control sequence for crRNA 5 target	negative control for CRISPR-LbCas12a system	28
BBa_K2927025	Negative control sequence for crRNA 6 target	negative control for CRISPR-LbCas12a system	28
BBa_K2927031	Positive control sequence for crRNAs target	positive control for CRISPR-LbCas12a system	355
BBa_K2927051	pHMT-LbCas12a plasmid (LbCas12a protein purification)	Produce 10X His-MBP-TEV site-LbCas12a proteins (connection)	4969
BBa_K2927061	T7 Pro-guide RNA- crRNA 1- crRNA 2 mutation	For the production of crRNA and negative control for CRISPR-LbCas12a system	106
BBa_K2927062	T7 Pro-guide RNA- crRNA 2- crRNA 1 mutation	For the production of crRNA and negative control for CRISPR-LbCas12a system	106
BBa_K2927063	T7 Pro-guide RNA- crRNA 3- crRNA 6 mutation	For the production of crRNA and negative control for CRISPR-LbCas12a system	106
BBa_K2927064	T7 Pro-guide RNA- crRNA 4- crRNA 5 mutation	For the production of crRNA and negative control for CRISPR-LbCas12a system	106
BBa_K2927065	T7 Pro-guide RNA- crRNA 5- crRNA 4 mutation	For the production of crRNA and negative control for CRISPR-LbCas12a system	106
BBa_K2927066	T7 Pro-guide RNA- crRNA 6- crRNA 3 mutation	For the production of crRNA and negative control for CRISPR-LbCas12a system	106

Basic Parts

Part number	Description	Supporting information	Length (bp)
BBa_K1614000	T7 Promoter	RNA polymerase binding	18
BBa_K2927001	RBS	Ribosome binding site	23
BBa_K2927002	10X His	For Ni2+ column	30
BBa_K2927003	MBP (maltose binding protein)	For MBP trap column	1101
BBa_K2927004	Tobaco Etch Virus (TEV) motif	TEV protein cutting site	21
BBa_K2927005	LbCas12a	Produce LbCas12a protein	3684
BBa_K395601	T7 Terminator	RNA polymerase leaving	48
BBa_K2927006	guide RNA	guide LbCas12a protein for binding target sequence	12
BBa_K2927010	DNA template of crRNA 1 for LbCas12a	DNA of ASFV for the detection	24
BBa_K2927011	DNA template of crRNA 2 for LbCas12a	DNA of ASFV for the detection	24
BBa_K2927012	DNA template of crRNA 3 for LbCas12a	DNA of ASFV for the detection	24
BBa_K2927013	DNA template of crRNA 4 for LbCas12a	DNA of ASFV for the detection	24
BBa_K2927014	DNA template of crRNA 5 for LbCas12a	DNA of ASFV for the detection	24
BBa_K2927015	DNA template of crRNA 6 for LbCas12a	DNA of ASFV for the detection	24
BBa_K2927020	Negative control sequence for crRNA 1 target	negative control for CRISPR-LbCas12a system	28
BBa_K2927021	Negative control sequence for crRNA 2 target	negative control for CRISPR-LbCas12a system	28
BBa_K2927022	Negative control sequence for crRNA 3 target	negative control for CRISPR-LbCas12a system	28
BBa_K2927023	Negative control sequence for crRNA 4 target	negative control for CRISPR-LbCas12a system	28
BBa_K2927024	Negative control sequence for crRNA 5 target	negative control for CRISPR-LbCas12a system	28
BBa_K2927025	Negative control sequence for crRNA 6 target	negative control for CRISPR-LbCas12a system	28
BBa_K2927031	Positive control sequence for crRNAs target	positive control for CRISPR-LbCas12a system	355

Composite Parts

Part number	Description	Supporting information	Length (bp)
BBa_K2927051	pHMT-LbCas12a plasmid (LbCas12a protein purification)	Produce 10X His-MBP-TEV site-LbCas12a proteins (connection)	4969
BBa_K2927061	T7 Pro-guide RNA- crRNA 1- crRNA 2 mutation	For the production of crRNA and negative control for CRISPR-LbCas12a system	106
BBa_K2927062	T7 Pro-guide RNA- crRNA 2- crRNA 1 mutation	For the production of crRNA and negative control for CRISPR-LbCas12a system	106
BBa_K2927063	T7 Pro-guide RNA- crRNA 3- crRNA 6 mutation	For the production of crRNA and negative control for CRISPR-LbCas12a system	106
BBa_K2927064	T7 Pro-guide RNA- crRNA 4- crRNA 5 mutation	For the production of crRNA and negative control for CRISPR-LbCas12a system	106
BBa_K2927065	T7 Pro-guide RNA- crRNA 5- crRNA 4 mutation	For the production of crRNA and negative control for CRISPR-LbCas12a system	106
BBa_K2927066	T7 Pro-guide RNA- crRNA 6- crRNA 3 mutation	For the production of crRNA and negative control for CRISPR-LbCas12a system	106

References

1. Chen, J. S., Ma, E., Harrington, L. B., Da Costa, M., Tian, X., Palefsky, J. M., & Doudna, J. A. (2018). CRISPR-Cas12a target binding unleashes indiscriminate single-stranded DNase activity. Science, 360(6387), 436-439.

2. Vlasova,N.N et al. (2014, Apr 06) African swine fever virus p72 (B646L) gene, complete cds.Retrieved from https://www.ncbi.nlm.nih.gov/nucleotide/KJ195685.1?report=genbank&log$=nucltop&blast_rank=10&RID=CP56X3A1014

3. NEW ENGLAND BioLabs. How do I design a guide RNA for use with EnGen Lba Cas12a? Retrieved from https://international.neb.com/faqs/2018/05/03/how-do-i-design-a-guide-rna-for-use-with-engen-lba-cas12a