Synthetic Biology Journal ›› 2023, Vol. 4 ›› Issue (4): 703-719.DOI: 10.12211/2096-8280.2022-054
• Invited Review • Previous Articles Next Articles
Mengdan MA1,2,3, Mengyu SHANG1,2,4, Yuchen LIU1,2
Received:
2022-09-30
Revised:
2023-02-01
Online:
2023-09-14
Published:
2023-08-31
Contact:
Yuchen LIU
马孟丹1,2,3, 尚梦宇1,2,4, 刘宇辰1,2
通讯作者:
刘宇辰
作者简介:
基金资助:
CLC Number:
Mengdan MA, Mengyu SHANG, Yuchen LIU. Application and prospect of CRISPR-Cas9 system in tumor biology[J]. Synthetic Biology Journal, 2023, 4(4): 703-719.
马孟丹, 尚梦宇, 刘宇辰. CRISPR-Cas9系统在肿瘤生物学中的应用及前景[J]. 合成生物学, 2023, 4(4): 703-719.
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URL: https://synbioj.cip.com.cn/EN/10.12211/2096-8280.2022-054
Fig. 2 Design and construction of the AND gate genetic circuits[50](a) The hUPⅡ and hTERT promoters are the inputs to the circuit. The hUPⅡ promoter drives the transcription of Cas9 mRNA and the hTERT promoter is linked to the transcription of sgRNA targeting LacI. The output gene is driven by a LacI-controlled CMV promoter. The effector can be expressed only when both Cas9 protein and sgRNA are presented. (b)~(e) are schematic representations of the synthetic circuits (SC-1, SC-2, SC-3 and SC-4). The output genes were hRluc, hBAX, p21 and E-cadherin.
Fig. 3 Design and construction of the AND gate minigene circuits[51]The UPⅡ promoter drove the transcription of Cas9 mRNA, while the hTERT promoter was used to promote the transcription of sgRNA targeting LacI. The output Renilla luciferase gene was regulated by a LacI-controlled CMV promoter. The luciferase was expressed only when both UPⅡ promoter and TERT promoter were both activated. In the design of the minigene circuit, the UPⅡ and hTERT promoters were replaced by their respective transcription factor binding elements. Both c-Myc and Get1, only in bladder cancer cells, had a relative high expression level at the same time. After initial expression of sgRNA1 and sgRNA2, they could further bind upstream of their own transcription initiation sites and then amplify the transcription signals of c-Myc and Get1 through the positive feedback mechanism to amplify their corresponding downstream genes transcription, respectively. Furthermore, the LacI gene was knocked out by sgRNA2, and luciferase reporter gene was activated by transcription. In normal bladder epithelial cells, luciferase could not be effectively transcribed and was further silenced by trace amounts of LacI expressed at the background level
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