In this issue, we introduce the Hemophilia B disease mouse model - F9 Knockout (KO) Mice. Hemophilia is a group of hereditary bleeding disorders characterized by a deficiency in clotting factor activity, leading to prolonged clotting time and a risk of bleeding even after minor injuries. Patients with severe forms of the disease can experience "spontaneous" bleeding without any apparent injury. As an X-linked recessive congenital rare disease, hemophilia predominantly affects males, with approximately 1 in every 30,000 newborn males worldwide diagnosed with Hemophilia B.[1]

Hemophilia B is Caused by F9 Gene Mutations

Currently, the most prevalent types of hemophilia are Hemophilia A and Hemophilia B. Hemophilia A patients suffer from a deficiency of Coagulation Factor VIII (factor 8) due to mutations in the F8 gene, while Hemophilia B results from mutations in the F9 gene leading to a lack of Coagulation Factor IX (factor 9). The F9 gene encodes a vitamin K-dependent serine protease, Coagulation Factor IX, which plays a crucial role in the intrinsic blood clotting pathway. Factor IX circulates in the blood as an inactive zymogen and is converted into its active form, IXa, through the cleavage by another coagulation factor called Factor XIa. In the coagulation cascade, Factor IXa activates Factor X through interactions with Ca2+ ions, membrane phospholipids, and Factor VIIIa (illustrated in Figure 1). Normal hemostasis can be achieved when the levels of Factors VIII and IX are ≥ 50% of the normal values.[2] A deficiency in the F9 gene can lead to a coagulation disorder due to insufficient Factor IX, causing Hemophilia B, an X-linked recessive inherited bleeding disorder.

Figure 1. Schematic Diagram of the Coagulation Pathway [2]

The severity of Hemophilia B is typically related to the level of Factor IX activity in the plasma

  • Patients with mild Hemophilia B (Factor IX activity >5%, >0.05 IU/mL) do not experience spontaneous bleeding but may have increased bleeding after trauma or surgery;
  • Those with moderate Hemophilia B (Factor IX activity 1%-5%, 0.01-0.05 IU/mL) rarely experience spontaneous bleeding, but even minor injuries can cause prolonged bleeding;
  • Severe Hemophilia B patients (Factor IX activity <1%, <0.01 IU/mL) may suffer from spontaneous bleeding, bleeding into soft tissues or joints, and severe subcutaneous hematomas.[3]

According to the Centers for Disease Control and Prevention (CDC), patients with severe Hemophilia B account for 30%-40% of all diagnosed cases of Hemophilia B.

Construction of F9 Gene Knockout Model

Cyagen has utilized gene editing technology to knockout (KO) exons 1-8 of the mouse F9 gene, constructing the F9 KO mouse model (product number: C001509). This model exhibits coagulation dysfunction and other Hemophilia B-related phenotypes, making it suitable for studying the genetic mechanisms and clinical phenotypes of human Hemophilia B. The F9 KO mouse model can aid in the development, screening, and evaluation of therapeutic drugs for Hemophilia B.

F9 KO Model Validation Data

(1) Absence of F9 mRNA expression in F9 KO mice

Figure 2. Detection of F9 mRNA Expression in F9 KO Mice and Wild-Type Mice (WT)

RT-qPCR results indicate that there is no expression of F9 mRNA in the liver and gallbladder of F9 KO mice, confirming the successful construction of the gene-edited model.

(2) Abnormalities in the four coagulation parameters of F9 KO mice suggest the presence of coagulation dysfunction in the mice.

Figure 3. Coagulation Parameters of F9 KO Mice and Wild-Type Mice (WT)

The results show that compared to wild-type mice, F9 KO mice have significantly prolonged activated partial thromboplastin time (APTT), indicating coagulation dysfunction and extended bleeding time, similar to the phenotype of the classic F9 KO disease model.[5] Furthermore, there are no significant differences in prothrombin time (PT), thrombin time (TT), and fibrinogen (FIB) between F9 KO mice and wild-type mice, aligning with the characteristics of coagulation parameter changes in clinical Hemophilia B patients.[6]

Cyagen Hematopoietic Disease Models: Mice, Rats, and more!

In addition to Hemophilia B, Cyagen has developed a range of genetically modified models for preclinical hematology research across a range of hematopoietic diseases – especially rare blood disorders  including gene knockouts (KOs), gene knock-ins (KIs), and point mutations (PMs). Furthermore, we can customize or collaboratively develop models based on your research requirements, such as developing mutation models with relevance for human disease pathogenicity. If you are interested in our blood disorder related models or wish to collaborate with us to develop models that meet your research needs, please contact us today for a free consultation and quote!

Target Gene Related Disease Product Number Product Name
F8 Hemophilia A C001211 F8 KO
F9 Hemophilia B C001509 F9 KO
Hbb Beta-thalassemia (HBB) C001508 Hbb-bs&Hbb-bt DKO
Flvcr1 Congenital Dyserythropoietic Anemia S-CKO-06994 C57BL/6J-Flvcr1em1Cflox/Cya
Flvcr1 Congenital Dyserythropoietic Anemia S-KO-06025 C57BL/6J-Flvcr1em1C/Cya
Usp1 Fanconi Anemia S-CKO-07336 C57BL/6J-Usp1em1Cflox/Cya
Usp1 Fanconi Anemia S-KO-06331 C57BL/6J-Usp1em1C/Cya

[1] Goodeve AC. Hemophilia B: molecular pathogenesis and mutation analysis. J Thromb Haemost. 2015 Jul;13(7):1184-95.
[2] Merck Manuals Professional Edition. Hemophilia - Hematology and Oncology - MSD Manual Professional Edition. [Online] Available at: [Accessed 12 Dec. 2023].
[3] Soroka AB, Feoktistova SG, Mityaeva ON, Volchkov PY. Gene Therapy Approaches for the Treatment of Hemophilia B. International Journal of Molecular Sciences. 2023; 24(13):10766.
[4] Centers for Disease Control and Prevention. Community Counts Registry Report – Males with Hemophilia 2014–2017. [Online] Available at:
[5] Wang L, Zoppè M, Hackeng TM, Griffin JH, Lee KF, Verma IM. A factor IX-deficient mouse model for hemophilia B gene therapy. Proc Natl Acad Sci U S A. 1997 Oct 14;94(21):11563-6.
[6] Thrombosis and Hemostasis Group, Hematology Society of Chinese Medical Association; Hemophilia Treatment Center Collaborative Network of China. [Consensus of Chinese expert on the diagnosis and treatment of hemophilia (version 2017)]. Zhonghua Xue Ye Xue Za Zhi. 2017 May 14;38(5):364-370. Chinese.